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Sample records for acids lysine methionine

  1. Amino acid nutrition beyond methionine and lysine for milk protein

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Amino acids are involved in many important physiological processes affecting the production, health, and reproduction of high-producing dairy cows. Most research and recommendations for lactating dairy cows has focused on methionine and lysine for increasing milk protein yield. This is because these...

  2. Short communication: Supplementing lysine and methionine in a lactation diet containing a high concentration of wet corn gluten feed did not alter milk protein yield.

    PubMed

    Mullins, C R; Weber, D; Block, E; Smith, J F; Brouk, M J; Bradford, B J

    2013-08-01

    Primiparous (n=33) and multiparous (n=63) lactating Holstein cows (186±51 d in milk) were used to evaluate the effects of supplementing metabolizable amino acids using lysine in a matrix of Ca salts of fatty acids (Megamine-L, Arm & Hammer Animal Nutrition, Princeton, NJ) and the isopropyl ester of 2-hydroxy-4-(methylthio) butanoic acid (MetaSmart, Adisseo Inc., Antony, France) in diets containing >26% wet corn gluten feed (dry matter basis). Cows were blocked by production level, parity, and pregnancy status, then randomly assigned to 1 of 8 pens and allowed a 7-d adaption period before receiving dietary treatments for 28 d. Pens were assigned randomly to either of 2 diets formulated to differ by metabolizable amino acid supply. Dry matter intake and production were monitored daily and milk components analyzed 3d/wk. Data were analyzed using mixed models with repeated measures. The original design of the study consisted of a control diet predicted to be deficient in lysine and methionine; however, after ingredient nutrients were analyzed and modeled with animal requirements at dry matter intake [26.6±0.35 kg/d (mean ± SEM)] and milk production levels achieved during the study (40.1±0.46 kg/d), only marginal deficiencies were predicted for the control (-8.1g/d for lysine; -1g/d for methionine) according to the National Research Council method, whereas the Cornell Net Carbohydrate and Protein System 5.0 and 6.1 models indicated positive balances for these amino acids (25.9 and 21.8 g/d for lysine, 14.7 and 18.9 g/d for methionine, respectively). Supplementing 30 g/d of metabolizable lysine in a Ca soap matrix and 2.4 g/d of metabolizable methionine as 2-hydroxy-4-(methylthio) butanoic acid led to positive predicted lysine and methionine balances by all 3 models, and predicted metabolizable lysine-to-methionine ratios ranging from 2.9 to 3.1. No treatment effects were observed for dry matter intake, milk yield, milk component concentrations or yields, or energy

  3. Influence of methionine supplementation of growing diets enriched with lysine on feedlot performance and characteristics of digestion in Holstein steer calves

    PubMed Central

    Torrentera, Noemí; Carrasco, Ramsés; Salinas-Chavira, Jaime; Plascencia, Alejandro; Zinn, Richard A.

    2017-01-01

    Objective Two trials were conducted in order to examine the effects of level of supplemental methionine on productive performance, dietary energetic, plasma amino acid concentration, and digestive function. Methods Dietary treatments consisted of a steam-flaked corn-based diet containing urea as the only source of supplemental nitrogen supplemented with no supplemental amino acid (control), or control plus 1.01% lysine and 0.032%, 0.064%, 0.096%, or 0.128% methionine. In Trial 1, 150 Holstein steer calves (127±4.9 kg) were utilized to evaluate the influence of treatments on growth-performance, dietary energetic, plasma amino acid concentration during the first 112 days of growing period. During the initial 56-d period calves received the 5 experimental diets. During the subsequent 56-d period all calves were fed the control diet. Results During the initial 56-d period, methionine supplementation increased (linear effect, p<0.01) plasma methionine. In the presence of supplemental lysine, increases on level of methionine in diet did not affect average daily gain. However, increased gain efficiency (quadratic effect, p = 0.03) and estimated dietary net energy (NE; linear effect, p = 0.05). Estimated metabolizable methionine supply was closely associated (R2 = 0.95) with efficiency NE utilization for maintenance and gain. During the subsequent 56-d period, when all calves received the control diet (no amino acid supplementation), plasma amino acid concentrations and growth performance was not different among groups. However, the effects of methionine supplementation during the initial 56-period carried over, so that following a 56-d withdrawal of supplementation, the overall 112-d effects on gain efficiency (quadratic effect, p = 0.05) dietary NE (linear effect, p≤0.05) remained appreciable. In Trial 2, 5 cannulated Holstein steers were used to evaluate treatment effects on characteristics of digestion and amino acid supply to the small intestine. There were no

  4. Rumen-protected methionine and lysine: effects on milk production and plasma amino acids of dairy cows with reference to metabolisable protein status.

    PubMed

    Awawdeh, Mofleh S

    2016-05-01

    Two experiments were conducted to study the effects of rumen-protected Met (RPM) alone or with rumen-protected Lys (RPL) on milk yield and plasma amino acids of dairy cows. In experiment 1, 24 multiparous Holstein cows (154 DIM) were assigned to one of 3 groups where each cow received 0 g/d of RPM and RPL (C), 30 g/d of RPM (M), or 30 g/d of RPM plus 25 g of RPL (ML). The study lasted for 8 weeks where milk yield and composition were determined weekly. Daily milk yield averaged 28·0, 27·8, and 29·7 kg/cow for the C, M, and ML groups, respectively. Dietary treatments had no effects (P ≥ 0·54) on milk contents of fat, lactose, solid non-fat or total solids. Milk protein content in the ML group was greater (P < 0·05) than the C and M groups. Plasma levels of all AA were not significantly (P ≥ 0·09) affected by supplemental RPL and/or RPM. In experiment 2, 30 multiparous Holstein cows (100 DIM) were assigned to one of 3 groups where each cow received 0 g/d of RPM and RPL (C), 50 g/d of RPM (M), or 50 g/d of RPM plus 25 g/d of RPL (ML). The study lasted for 5 weeks. Cows in the M (30·5 kg) and ML (31·4 kg) groups produced (P < 0·05) more milk than those of the C group (29·1 kg). Under conditions of this study, RPM plus RPL improved milk yield and protein contents of dairy cows and was better than supplying RPM alone. Response in milk yield to RPM and RPL was affected by the MP status of cows which deserves further investigation.

  5. Adipic acid increases plasma lysine but does not improve the efficiency of lysine utilization in swine.

    PubMed

    van Kempen, T A; van Heugten, E; Trottier, N L

    2001-09-01

    Adipic acid, upon catabolism, results in intermediates that bear a structural similarity to lysine degradation products. The objectives of this research were to determine whether adipic acid affects lysine concentrations in plasma and to evaluate whether adipic acid improves the efficiency of lysine utilization in pigs. In Exp. 1, nursery pigs (n = 14) were fed (for a period of 7 d) either a standard nursery diet or the same diet supplemented with 1% adipic acid to assess effects on plasma amino acid concentrations (plasma collected on d 7). In Exp. 2, nursery pigs (n = 56) were fed (for a period of 15 d) either a control diet or the same diet but deficient in either lysine, threonine, or tryptophan with or without supplemental adipic acid to assess the effects of adipic acid on the efficiency of amino acid utilization. The results from Exp. 1 showed that adipic acid increased plasma lysine (by 18%) but not alpha-amino adipic acid, an intermediate in lysine degradation. Experiment 2 demonstrated that adipic acid did not increase the efficiency of utilization of lysine, threonine, or tryptophan. The lack of effects on alpha-amino adipic acid in Exp. 1 and the lack of a positive effect on the efficiency of utilization of lysine, threonine, and tryptophan suggest that adipic acid does not inhibit the mitochondrial uptake of lysine and(or) its degradation in the mitochondrion. It is concluded that feeding adipic acid increases plasma lysine but does not improve the efficiency of lysine utilization.

  6. Kinetics of the reactions of hypochlorous acid and amino acid chloramines with thiols, methionine, and ascorbate.

    PubMed

    Peskin, A V; Winterbourn, C C

    2001-03-01

    Thiol oxidation by hypochlorous acid and chloramines is a favorable reaction and may be responsible for alterations in regulatory or signaling pathways in cells exposed to neutrophil oxidants. In order to establish the mechanism for such changes, it is necessary to appreciate whether these oxidants are selective for different thiols as compared with other scavengers. We have measured rate constants for reactions of amino acid chloramines with a range of thiols, methionine, and ascorbate, using a combination of stopped-flow and competitive kinetics. For HOCl, rate constants are too fast to measure directly by our system and values relative to reduced glutathione were determined by competition with methionine. For taurine chloramine, the rate constants for reaction with 5-thio-2-nitrobenzoic acid, GSH, methionine, and ascorbate at pH 7.4 were 970, 115, 39, and 13 M(-1) s(-1), respectively. Values for 10 thiols varied by a factor of 20 and showed an inverse relationship to the pK(a) of the thiol group. Rate constants for chloramines of glycine and N-alpha-acetyl-lysine also showed these relationships. Rates increased with decreasing pH, suggesting a mechanism involving acid catalysis. For hypochlorous acid, rates of reaction with 5-thio-2-nitrobenzoic acid, GSH, cysteine, and most of the other thiols were very similar. Relative reactivities varied by less than 5 and there was no dependence on thiol pK(a). Chloramines have the potential to be selective for different cellular thiols depending on their pK(a). For HOCl to be selective, other factors must be important, or its reactions could be secondary to chloramine formation.

  7. S-adenosyl methionine is necessary for inhibition of the methyltransferase G9a by the lysine 9 to methionine mutation on histone H3.

    PubMed

    Jayaram, Hariharan; Hoelper, Dominik; Jain, Siddhant U; Cantone, Nico; Lundgren, Stefan M; Poy, Florence; Allis, C David; Cummings, Richard; Bellon, Steven; Lewis, Peter W

    2016-05-31

    Lysine to methionine (K-to-M) mutations in genes encoding histone H3 are thought to drive a subset of pediatric brain and bone cancers. These high-frequency K-to-M mutations occur at sites of methylation on histone H3, and tumors containing the mutant histones exhibit a global loss of specific histone methylation marks. Previous studies showed that K-to-M mutant histones, also known as oncohistones, are potent orthosteric inhibitors of specific Su(var)3-9, Enhancer-of-zeste, Trithorax (SET) domain methyltransferases. However, the biochemical and biophysical details of the interaction between K-to-M mutant histones and the respective SET domain methyltransferases are currently unknown. Here, we use the histone H3K9-directed methyltransferase G9a as a model to explore the mechanism of inhibition by K-to-M oncohistones. X-ray cocrystal structures revealed that the K9M residue of histone H3 occupies the active site cavity of G9a, and kinetic analysis indicates competitive inhibition of G9a by histone H3K9M. Additionally, we find that the cofactor S-adenosyl methionine (SAM) is necessary for stable interaction between G9a and H3K9M histone. Consistent with the formation of a ternary complex, we find that the inhibitory peptide is uncompetitive with regard to SAM. These data and others indicate that K-to-M oncohistones promote global loss of specific lysine methylation through sequestration and inhibition of SAM-bound SET domain methyltransferases.

  8. Comparative analysis of some essential amino acids and available lysine in Acacia colei and A. tumida seeds using chemical methods and an amino acid analyzer.

    PubMed

    Falade, Olumuyiwa S; Adewusi, Steve R A

    2013-01-01

    Methionine, cysteine, tryptophan, and available lysine were determined in Acacia colei and A. tumida seeds and some cereals using chemical methods, and the results were compared to those obtained using an amino acid analyzer. Ba(OH)2 hydrolysis gave the best result of the three methods of hydrolysis (acid, base, and enzyme) tried. Oxidized methionine, cysteine, and tryptophan were not detected, but S-carboxyethylcysteine was estimated as cysteine by the chemical methods, thus overestimating cysteine's content in Acacia seeds. Tryptophan and methionine were higher in cereals than in Acacia seeds, while the level of cysteine and available lysine was higher in Acacia seeds than in cereals. These results agreed with values obtained using the amino acid analyzer and could therefore be used in low budget laboratories.

  9. Source of carbohydrate and metabolizable lysine and methionine in the diet of recently weaned dairy calves on digestion and growth.

    PubMed

    Hill, T M; Quigley, J D; Bateman, H G; Aldrich, J M; Schlotterbeck, R L

    2016-04-01

    Two 56-d trials with weaned Holstein dairy calves (initially 72 ± 1.8 kg of body weight, 58 to 60 d of age) fed 95% concentrate and 5% chopped grass hay diets were conducted. Each trial used 96 calves (4 calves/pen). During 15 of the last 21 d of the first trial and 10 of 14 d of the second and third week of the second trial, fecal samples were taken to estimate digestibility using acid-insoluble ash as an internal marker. Digestibility estimates along with 56-d average daily gain (ADG), hip width change, body condition score, and fecal score were analyzed with pen as the experimental unit. In trial 1, a textured diet (19% crude protein) with high starch [52% starch, 13% neutral detergent fiber (NDF)] based on whole corn and oats or a pelleted low-starch (20% starch, 35% NDF), high-digestible fiber diet were used. Within starch level, diets were formulated from supplemental soybean meal or soybean meal with blood meal and Alimet (Novus International Inc., St. Charles, MO) to provide 2 metabolizable protein levels (1 and 1.07% metabolizable lysine plus methionine). The 4 treatments were analyzed as a completely randomized design with a 2 by 2 factorial arrangement (6 pens/diet). In trial 2, all pelleted diets (19% crude protein) were fed. Diets were based on soybean hulls, wheat middlings, or corn, which contained increasing concentrations of starch (13, 27, and 42% starch and 42, 23, and 16% NDF, respectively; 8 pens/diet). Contrast statements were constructed to separate differences in the means (soybean hulls plus wheat middlings vs. corn; soybean hulls vs. wheat middlings). In trial 1, intake of organic matter (OM) did not differ. Digestibility of OM was greater in calves fed high- versus low starch-diets. Digestibility of NDF and starch were less in calves fed the high- versus low-starch diets. Calf ADG and hip width change were greater for high- versus low-starch diets. Source of protein did not influence digestibility or ADG. In trial 2, intake of OM was not

  10. Effects of castration age, dietary protein level and lysine/methionine ratio on animal performance, carcass and meat quality of Friesian steers intensively reared.

    PubMed

    Prado, I N; Campo, M M; Muela, E; Valero, M V; Catalan, O; Olleta, J L; Sañudo, C

    2014-09-01

    The effects of castration age, dietary protein level and the dietary lysine/methionine (lys/met) ratio on animal performance, carcass characteristics and meat quality were studied in 64 intensively reared Friesian steers. Animals underwent castration procedures at 15 days old or at 5 months old. Dietary treatments started at 90 days old, with eight animals from each castration age randomly allocated to each treatment: 14.6% v. 16.8% CP (DM basis), and 3.0 v. 3.4 lys/met, on a 2×2×2 design. The recommended ratio of 3.0 was reached with supplementation of protected methionine. Steers were slaughtered at 443.5±26.2 kg live weight when they reached 12 months old approximately. Average daily gain, cold carcass weight or carcass classification were not affected by any studied effect. Muscle moisture (P=0.024), C18:2n-6 percentage (P=0.047), polyunsaturated fatty acid/saturated fatty acid (P=0.049) and n-6/n-3 (P=0.003) were higher in late castrated animals. Both high levels of dietary protein (P=0.008) and lys/met ratio (P=0.048) increased the percentage of muscle in the carcass. A level of 16.8% of CP in the diet also increased the percentage of monounsaturated fatty acids in the intramuscular fat (P=0.032), whereas a ratio lys/met of 3.4 decreased the percentage of saturated fatty acids (P=0.028). Thus, it is recommended using diets with a high protein level (16.8%) and a high lys/met ratio (3.4) in animals slaughtered at a young age, in order to obtain carcasses with high muscle content without negatively affecting productive traits or intramuscular fat composition.

  11. Removal of N-terminal methionine from haemoglobin nascent peptides by a membrane-bound rat liver methionine aminopeptidase.

    PubMed Central

    Termignoni, C; Freitas, J O; Guimarães, J A

    1986-01-01

    A membrane-bound aminopeptidase able to remove methionine from haemoglobin nascent peptides is described. The enzyme also hydrolyses methionine from methionyl-lysyl-bradykinin but not lysine from lysyl-bradykinin. The tripeptide Met-Ala-Ser is poorly hydrolysed. This aminopeptidase also splits amino acid 2-naphthylamides, being, however, less specific with respect to these synthetic substrates. PMID:3087345

  12. Divergent modulation of swine ileal microbiota by formic acid and methionine hydroxy analogue-free acid.

    PubMed

    Apajalahti, J; Rademacher, M; Htoo, J K; Redshaw, M; Kettunen, A

    2009-06-01

    Management of intestinal microbiota of monogastric animals has increased in importance since the ban of growth promoting antibiotics in many countries. Organic acids have been used as alternatives to antibiotics by many feed manufacturers. Regardless of the wide usage, the effect, dose response and mode of action of acids on intestinal microbes is poorly understood. In this study, we investigated the effects of dietary supplementation of three commonly used products, namely formic acid (FA) (90%), dl-methionine (DLM) (99%) and liquid methionine hydroxy analogue-free acid (88%), on ileal microbiota of pigs. Laboratory simulation system, mimicking swine ileum, was used to study the products at various concentrations and combinations. Furthermore, selected combinations were tested in a piglet trial to confirm the findings made in in vitro studies. FA turned out to have a dual effect on ileal microbiota. At concentrations below 0.5%, it significantly stimulated bacteria, but at higher inclusion rates it was highly inhibitory. This finding, which was consistent in in vitro and in vivo studies, implies that reducing the dose of FA does not lead to a diluted inhibitory effect, but in fact, an opposite, stimulatory effect on intestinal microbiota. It is highly important that feed compounders acknowledge this finding. Unlike FA, the inhibitory effect of methionine hydroxy analogue on ileal bacteria was linearly dose dependent and significant at inclusion levels above 0.2%, in vitro. Partial replacement of methionine hydroxy analogue by FA, or FA by methionine hydroxy analogue, led to an unpredictable outcome due to the dual effects of FA; e.g., a minor inclusion of added FA changed the inhibitory effect of methionine hydroxy analogue into microbial stimulation by FA. Inhibition of ileal microbiota by methionine hydroxy analogue was detected only in in vitro studies, suggesting that intact methionine hydroxy analogue may not have reached the ileum, in live animals. Therefore

  13. Lysine, methionine, phenylalanine, arginine, valine, isoleucine, leucine, and threonine maintenance requirements of broiler breeders.

    PubMed

    Sakomura, N K; Ekmay, R D; Mei, S J; Coon, C N

    2015-11-01

    Five hundred and sixty Cobb-Vantress broiler breeders were used to determine the maintenance requirement of dietary Arg, His, Ile, Leu, Lys, Met, Phe, Thr, and Val in 10 21-d assays using the comparative slaughter technique. Fifty sexually immature broiler breeder pullets per assay were given crystalline amino acid diets containing graded levels of Arg, His, Ile, Leu, Lys, Met, Phe, Thr, or Val, respectively, representing 0, 10, 20, 30, 40% of their suggested requirement level (NRC, 1994) with all other amino acids maintained at 40% of their suggested requirement level. Sixty hens were slaughtered prior to the beginning of the study to assess initial body composition at 3 weight groups. Linear regression lines (protein accretion vs. amino acid intake) were determined and the maintenance requirements were calculated based on zero protein accretion. The maintenance requirements expressed on a metabolic weight basis were determined to be 174, 94, 52, 81, 60, 126, 133, and 155 mg/kg(0.75)/d for Arg, Ile, Leu, Lys, Met, Phe, Thr, and Val, respectively. Additionally, the maintenance requirements, expressed on metabolic protein weight basis, were determined to be 651, 329, 172, 295, 223, 523, 478, and 546 mg/kgCP/d for Arg, Ile, Leu, Lys, Met, Phe, Thr, and Val, respectively. The requirement for His could not be established as the results obtained were outside of assay range.

  14. Myeloperoxidase-mediated protein lysine oxidation generates 2-aminoadipic acid and lysine nitrile in vivo.

    PubMed

    Lin, Hongqiao; Levison, Bruce S; Buffa, Jennifer A; Huang, Ying; Fu, Xiaoming; Wang, Zeneng; Gogonea, Valentin; DiDonato, Joseph A; Hazen, Stanley L

    2017-03-01

    Recent studies reveal 2-aminoadipic acid (2-AAA) is both elevated in subjects at risk for diabetes and mechanistically linked to glucose homeostasis. Prior studies also suggest enrichment of protein-bound 2-AAA as an oxidative post-translational modification of lysyl residues in tissues associated with degenerative diseases of aging. While in vitro studies suggest redox active transition metals or myeloperoxidase (MPO) generated hypochlorous acid (HOCl) may produce protein-bound 2-AAA, the mechanism(s) responsible for generation of 2-AAA during inflammatory diseases are unknown. In initial studies we observed that traditional acid- or base-catalyzed protein hydrolysis methods previously employed to measure tissue 2-AAA can artificially generate protein-bound 2-AAA from an alternative potential lysine oxidative product, lysine nitrile (LysCN). Using a validated protease-based digestion method coupled with stable isotope dilution LC/MS/MS, we now report protein bound 2-AAA and LysCN are both formed by hypochlorous acid (HOCl) and the MPO/H2O2/Cl(-) system of leukocytes. At low molar ratio of oxidant to target protein N(ε)-lysine moiety, 2-AAA is formed via an initial N(ε)-monochloramine intermediate, which ultimately produces the more stable 2-AAA end-product via sequential generation of transient imine and semialdehyde intermediates. At higher oxidant to target protein N(ε)-lysine amine ratios, protein-bound LysCN is formed via initial generation of a lysine N(ε)-dichloramine intermediate. In studies employing MPO knockout mice and an acute inflammation model, we show that both free and protein-bound 2-AAA, and in lower yield, protein-bound LysCN, are formed by MPO in vivo during inflammation. Finally, both 2-AAA and to lesser extent LysCN are shown to be enriched in human aortic atherosclerotic plaque, a tissue known to harbor multiple MPO-catalyzed protein oxidation products. Collectively, these results show that MPO-mediated oxidation of protein lysyl

  15. Amino acid distribution and metabolism in pituitary adenomas using positron emission tomography with D-(/sup 11/C)methionine and L-(/sup 11/C)methionine

    SciTech Connect

    Bergstroem, M.M.; Muhr, C.; Lundberg, P.O.; Bergstroem, K.L.; Lundqvist, H.; Antoni, G.; Fasth, K.J.; Langstroem B3

    1987-05-01

    Four patients with hormonally inactive pituitary adenomas were examined with positron emission tomography (PET) after injection, during different examinations, of L-(methyl-/sup 11/C)methionine and D-(methyl-/sup 11/C)methionine, respectively. After the rapid distribution phase, the enantiomer L-(/sup 11/C)methionine, which is metabolically active, showed a considerable continuous irreversible trapping attributed to amino acid metabolism. The stereoisomer D-(/sup 11/C)methionine, which does not participate in protein synthesis, showed a rapid distribution within the whole adenoma tissue, with a distribution space on the order of 100%. A minimal irreversible trapping was observed which could be explained by technical factors. It is concluded that PET using the two enantiomers allows a separation of passive distribution and metabolism, and that L-(/sup 11/C)methionine can be used for in vivo quantitative studies of amino acid metabolism of pituitary adenomas.

  16. Effects of supplements of folic acid, vitamin B12, and rumen-protected methionine on whole body metabolism of methionine and glucose in lactating dairy cows.

    PubMed

    Preynat, A; Lapierre, H; Thivierge, M C; Palin, M F; Matte, J J; Desrochers, A; Girard, C L

    2009-02-01

    The present experiment was undertaken to determine the effects of dietary supplements of rumen-protected methionine and intramuscular injections of folic acid and vitamin B(12), given 3 wk before to 16 wk after calving, on glucose and methionine metabolism of lactating dairy cows. Twenty-four multiparous Holstein cows were assigned to 6 blocks of 4 cows each according to their previous milk production. Within each block, 2 cows were fed a diet estimated to supply methionine as 1.83% metabolizable protein, equivalent to 76% of methionine requirement, whereas the 2 other cows were fed the same diet supplemented daily with 18 g of rumen-protected methionine. Within each diet, the cows were administrated either no vitamin supplement or weekly intramuscular injections of 160 mg of folic acid plus 10 mg of vitamin B(12.) To investigate metabolic changes at 12 wk of lactation, glucose and methionine kinetics were measured by isotope dilution using infusions of 3[U-(13)C]glucose, [(13)C]NaHCO(3) and 3[1-(13)C,(2)H(3)] methionine. Milk and plasma concentrations of folic acid and vitamin B(12) increased with vitamin injections. Supplementary B-vitamins increased milk production from 34.7 to 38.9 +/- 1.0 kg/d and increased milk lactose, protein, and total solids yields. Whole-body glucose flux tended to increase with vitamin supplementation with a similar quantitative magnitude as the milk lactose yield increase. Vitamin supplementation increased methionine utilization for protein synthesis through increased protein turnover when methionine was deficient and through decreased methionine oxidation when rumen-protected methionine was fed. Vitamin supplementation decreased plasma concentrations of homocysteine independently of rumen-protected methionine feeding, although no effect of vitamin supplementation was measured on methionine remethylation, but this could be due to the limitation of the technique used. Therefore, the effects of these B-vitamins on lactation performance

  17. Effects of dietary lysine and methionine supplementation on ross 708 male broilers from 21 to 42 days of age (III): serum metabolites, hormones, and their relationship with growth performance

    Technology Transfer Automated Retrieval System (TEKTRAN)

    A previous study has shown that a limited increase of lysine (Lys) and methionine (Met) in broiler diets may improve feed conversion ratio (FCR), BW, carcass yield, and breast meat yield. This study was conducted to determine the effects of dietary Lys and Met supplementation on various blood serum ...

  18. Histidine-lysine peptides as carriers of nucleic acids.

    PubMed

    Leng, Qixin; Goldgeier, Lisa; Zhu, Jingsong; Cambell, Patricia; Ambulos, Nicholas; Mixson, A James

    2007-03-01

    With their biodegradability and diversity of permutations, peptides have significant potential as carriers of nucleic acids. This review will focus on the sequence and branching patterns of peptide carriers composed primarily of histidines and lysines. While lysines within peptides are important for binding to the negatively charged phosphates, histidines are critical for endosomal lysis enabling nucleic acids to reach the cytosol. Histidine-lysine (HK) polymers by either covalent or ionic bonds with liposomes augment transfection compared to liposome carriers alone. More recently, we have examined peptides as sole carriers of nucleic acids because of their intrinsic advantages compared to the bipartite HK/liposome carriers. With a protocol change and addition of a histidine-rich tail, HK peptides as sole carriers were more effective than liposomes alone in several cell lines. While four-branched polymers with a primary repeating sequence pattern of -HHK- were more effective as carriers of plasmids, eight-branched polymers with a sequence pattern of -HHHK- were more effective as carriers of siRNA. Compared to polyethylenimine, HK carriers of siRNA and plasmids had reduced toxicity. When injected intravenously, HK polymers in complex with plasmids encoding antiangiogenic proteins significantly decreased tumor growth. Furthermore, modification of HK polymers with polyethylene glycol and vascular-specific ligands increased specificity of the polyplex to the tumor by more than 40-fold. Together with further development and insight on the structure of HK polyplexes, HK peptides may prove to be useful as carriers of different forms of nucleic acids both in vitro and in vivo.

  19. Independent and additive effects of glutamic acid and methionine on yeast longevity.

    PubMed

    Wu, Ziyun; Song, Lixia; Liu, Shao Quan; Huang, Dejian

    2013-01-01

    It is established that glucose restriction extends yeast chronological and replicative lifespan, but little is known about the influence of amino acids on yeast lifespan, although some amino acids were reported to delay aging in rodents. Here we show that amino acid composition greatly alters yeast chronological lifespan. We found that non-essential amino acids (to yeast) methionine and glutamic acid had the most significant impact on yeast chronological lifespan extension, restriction of methionine and/or increase of glutamic acid led to longevity that was not the result of low acetic acid production and acidification in aging media. Remarkably, low methionine, high glutamic acid and glucose restriction additively and independently extended yeast lifespan, which could not be further extended by buffering the medium (pH 6.0). Our preliminary findings using yeasts with gene deletion demonstrate that glutamic acid addition, methionine and glucose restriction prompt yeast longevity through distinct mechanisms. This study may help to fill a gap in yeast model for the fast developing view that nutrient balance is a critical factor to extend lifespan.

  20. Independent and Additive Effects of Glutamic Acid and Methionine on Yeast Longevity

    PubMed Central

    Wu, Ziyun; Song, Lixia; Liu, Shao Quan; Huang, Dejian

    2013-01-01

    It is established that glucose restriction extends yeast chronological and replicative lifespan, but little is known about the influence of amino acids on yeast lifespan, although some amino acids were reported to delay aging in rodents. Here we show that amino acid composition greatly alters yeast chronological lifespan. We found that non-essential amino acids (to yeast) methionine and glutamic acid had the most significant impact on yeast chronological lifespan extension, restriction of methionine and/or increase of glutamic acid led to longevity that was not the result of low acetic acid production and acidification in aging media. Remarkably, low methionine, high glutamic acid and glucose restriction additively and independently extended yeast lifespan, which could not be further extended by buffering the medium (pH 6.0). Our preliminary findings using yeasts with gene deletion demonstrate that glutamic acid addition, methionine and glucose restriction prompt yeast longevity through distinct mechanisms. This study may help to fill a gap in yeast model for the fast developing view that nutrient balance is a critical factor to extend lifespan. PMID:24244480

  1. Methionine sulfoxide reductase contributes to meeting dietary methionine requirements

    PubMed Central

    Zhao, Hang; Kim, Geumsoo; Levine, Rodney L.

    2012-01-01

    Methionine sulfoxide reductases are present in all aerobic organisms. They contribute to antioxidant defenses by reducing methionine sulfoxide in proteins back to methionine. However, the actual in vivo roles of these reductases are not well defined. Since methionine is an essential amino acid in mammals, we hypothesized that methionine sulfoxide reductases may provide a portion of the dietary methionine requirement by recycling methionine sulfoxide. We used a classical bioassay, the growth of weanling mice fed diets varying in methionine, and applied it to mice genetically engineered to alter the levels of methionine sulfoxide reductase A or B1. Mice of all genotypes were growth retarded when raised on chow containing 0.10% methionine instead of the standard 0.45% methionine. Retardation was significantly greater in knockout mice lacking both reductases. We conclude that the methionine sulfoxide reductases can provide methionine for growth in mice with limited intake of methionine, such as may occur in the wild. PMID:22521563

  2. Sulfur amino acid deficiency upregulates intestinal methionine cycle activity and suppresses epithelial growth in neonatal pigs.

    Technology Transfer Automated Retrieval System (TEKTRAN)

    We recently showed that the developing gut is a significant site of methionine transmethylation to homocysteine and transsulfuration to cysteine. We hypothesized that sulfur amino acid (SAA) deficiency would preferentially reduce mucosal growth and antioxidant function in neonatal pigs. Neonatal pi...

  3. Pathways of Amino Acid Degradation in Nilaparvata lugens (Stål) with Special Reference to Lysine-Ketoglutarate Reductase/Saccharopine Dehydrogenase (LKR/SDH)

    PubMed Central

    Wan, Pin-Jun; Yuan, San-Yue; Tang, Yao-Hua; Li, Kai-Long; Yang, Lu; Fu, Qiang; Li, Guo-Qing

    2015-01-01

    Nilaparvata lugens harbors yeast-like symbionts (YLSs). In present paper, a genome-wide analysis found 115 genes from Ni. lugens and 90 genes from YLSs that were involved in the metabolic degradation of 20 proteinogenic amino acids. These 205 genes encoded for 77 enzymes. Accordingly, the degradation pathways for the 20 amino acids were manually constructed. It is postulated that Ni. lugens can independently degrade fourteen amino acids (threonine, alanine, glycine, serine, aspartate, asparagine, phenylalanine, tyrosine, glutamate, glutamine, proline, histidine, leucine and lysine). Ni. lugens and YLSs enzymes may work collaboratively to break down tryptophan, cysteine, arginine, isoleucine, methionine and valine. We cloned a lysine-ketoglutarate reductase/saccharopine dehydrogenase gene (Nllkr/sdh) that encoded a bifunctional enzyme catalyzing the first two steps of lysine catabolism. Nllkr/sdh is widely expressed in the first through fifth instar nymphs and adults, and is highly expressed in the fat body, ovary and gut in adults. Ingestion of dsNllkr/sdh by nymphs successfully knocked down the target gene, and caused nymphal/adult mortality, shortened nymphal development stage and reduced adult fresh weight. Moreover, Nllkr/sdh knockdown resulted in three defects: wings were shortened and thickened; cuticles were stretched and thinned; and old nymphal cuticles remained on the tips of legs and abdomen and were not completely shed. These data indicate that impaired lysine degradation negatively affects the survival and development of Ni. lugens. PMID:26000452

  4. Nonnatural amino acid incorporation into the methionine 214 position of the metzincin Pseudomonas aeruginosa alkaline protease

    PubMed Central

    Walasek, Paula; Honek, John F

    2005-01-01

    Background The alkaline protease from Pseudomonas aeruginosa (AprA) is a member of the metzincin superfamily of metalloendoproteases. A key feature of these proteases is a conserved methionine-containing 1,4-tight β turn at the base of the active site zinc binding region. Results To explore the invariant methionine position in this class of protease, incorporation of a nonnatural fluorinated methionine, L-difluoromethionine (DFM), into this site was accomplished. Although overproduction of the N-terminal catalytic fragment of AprA resulted in protein aggregates which could not be resolved, successful heterologous production of the entire AprA was accomplished in the presence and absence of the nonnatural amino acid. DFM incorporation was found to only slightly alter the enzyme kinetics of AprA. In addition, differential scanning calorimetry indicated no significant alteration in the thermal stability of the modified enzyme. Conclusion Although invariant in all metzincin proteases, the methionine 214 position in AprA can be successfully replaced by the nonnatural amino acid DFM resulting in little effect on protein structure and function. This study indicates that the increased size of the methyl group by the introduction of two fluorines is still sufficiently non-sterically demanding, and bodes well for the application of DFM to biophysical studies of protein structure and function in this class of protease. PMID:16221305

  5. Protective role of S-adenosyl-L-methionine against hydrochloric acid stress in Saccharomyces cerevisiae.

    PubMed

    Malakar, Dipankar; Dey, Anindya; Ghosh, Anil K

    2006-09-01

    S-adenosyl-L-methionine (AdoMet, 1mM) protects the stationary phase cells of Saccharomyces cerevisiae against the killing effect of acid (10mM HCl, pH approximately 2). Both the acid and the acid plus AdoMet treatment for 2h increased the plasma membrane H(+)-ATPase activity; thereafter it decreased to the basal level. AdoMet partially recovered the intracellular pH (pH(in)) that dropped in presence of acid. AdoMet treatment facilitated acid induced phospholipid biosynthesis as well as membrane proliferation, which was reflected in the cellular lipid composition.

  6. A study of the protein and amino acid requirements of the growing New Zealand White rabbit with emphasis on lysine and the sulphur-containing amino acids.

    PubMed

    Spreadbury, D

    1978-05-01

    1. New Zealand White (NZW) rabbits were given, between 4 and 8 weeks of age, a range of diets, based on oats and fish meal, containing from 104 to 255 g crude protein (nitrogen x 6.25; CP)/kg to establish the level of CP below which growth was retarded. 2. In three experiments each diet was fed to four animals and food intake, growth and N balance were measured over 4 weeks. Body analysis was also carried out after two of the experiments. 3. The rates of food intake and growth of animals increased with dietary CP concentration until a CP concentration of approximately 150 g/kg diet had been reached. Beyond this there was little further improvement. N balance studies showed that once this dietary concentration of CP had been reached, there was a reduced rate of N retention. 4. Good agreement was found between N retention measured by balance methods and by body analysis: body composition showed a tendency towards an increase 5. Microbial protein produced in the caecum and eaten during coprophagy, was found to supplement the dietary protein by approximately 2 g CP/d, or by only 0.1 of a normal dietary intake of CP. 6. In the second part of the study NZW rabbits were offered, between 5 and 8 weeks of age, diets based on oats containing 150 g CP/kg. The protein supplied by oats was supplemented with maize gluten, gelatin, groundnut meal, casein, soya-bean meal or fish meal. 7. Rabbits offered diets containing casein, soya-bean meal and fish meal gained 40-50 g/d similar, to animals given a well-balanced control diet, while those given diets containing maize gluten, gelatin or groundnut meal gained approximately 30 g/d. This indicated that amino acid balance in dietary protein was important to the growing rabbit. 8. In later experiments, diets based on cereals and groundnut meal supplemented with varying amounts of lysine and methionine were offered during a 3-week-post-weaning period in order to assess requirements for those limiting amino acids. 9. The addition of both

  7. Impact of food supplementation and methionine on high densities of cotton rats: Support of the amino-acid-quality hypothesis?

    USGS Publications Warehouse

    Webb, R.E.; Leslie, David M.; Lochmiller, R.L.; Masters, R.E.

    2005-01-01

    Considerable research supports the tenet that quantity and quality of food limit vertebrate populations. We evaluated predictions that increased availabilities of food and the essential amino acid methionine were related to population limitation of the hispid cotton rat (Sigmodon hispidus). Effects of supplemental food and methionine on density, survival, and reproductive parameters of wild cotton rats were assessed in north-central Oklahoma in 1998-1999. Twelve enclosed groups of 16 adult cotton rats each (8 male, 8 female) were randomly assigned to either no supplementation (control), supplementation with a mixed ration that had methionine at slightly below maintenance levels (0.20%), or a methionine-enhanced mixed ration (1.20%). In general, densities of cotton rats were twice as high and were sustained longer with dietary supplementation, and methionine-supplemented populations maintained the highest densities. Treatment effects on survival depended on time of year, with higher survival in supplemented enclosures in October and November. Per capita recruitment was highest with methionine-enhanced food. Treatment effects on proportions of overall and female cotton rats in reproductive condition depended on sampling date, but males were most reproductively active with methionine supplementation. Methionine supplementation resulted in an earlier and longer reproductive season. Density-dependent and density-independent factors no doubt interplay to determine population dynamics of cotton rats, but our results suggest that methionine plays a role in the population dynamics of wild cotton rats, apparently by enhancing overall density, recruitment, and reproductive activity of males.

  8. Comprehensive Profiling of Amino Acid Response Uncovers Unique Methionine-Deprived Response Dependent on Intact Creatine Biosynthesis

    PubMed Central

    Tang, Xiaohu; Keenan, Melissa M.; Wu, Jianli; Lin, Chih-An; Dubois, Laura; Thompson, J. Will; Freedland, Stephen J.; Murphy, Susan K.; Chi, Jen-Tsan

    2015-01-01

    Besides being building blocks for protein synthesis, amino acids serve a wide variety of cellular functions, including acting as metabolic intermediates for ATP generation and for redox homeostasis. Upon amino acid deprivation, free uncharged tRNAs trigger GCN2-ATF4 to mediate the well-characterized transcriptional amino acid response (AAR). However, it is not clear whether the deprivation of different individual amino acids triggers identical or distinct AARs. Here, we characterized the global transcriptional response upon deprivation of one amino acid at a time. With the exception of glycine, which was not required for the proliferation of MCF7 cells, we found that the deprivation of most amino acids triggered a shared transcriptional response that included the activation of ATF4, p53 and TXNIP. However, there was also significant heterogeneity among different individual AARs. The most dramatic transcriptional response was triggered by methionine deprivation, which activated an extensive and unique response in different cell types. We uncovered that the specific methionine-deprived transcriptional response required creatine biosynthesis. This dependency on creatine biosynthesis was caused by the consumption of S-Adenosyl-L-methionine (SAM) during creatine biosynthesis that helps to deplete SAM under methionine deprivation and reduces histone methylations. As such, the simultaneous deprivation of methionine and sources of creatine biosynthesis (either arginine or glycine) abolished the reduction of histone methylation and the methionine-specific transcriptional response. Arginine-derived ornithine was also required for the complete induction of the methionine-deprived specific gene response. Collectively, our data identify a previously unknown set of heterogeneous amino acid responses and reveal a distinct methionine-deprived transcriptional response that results from the crosstalk of arginine, glycine and methionine metabolism via arginine

  9. Comprehensive profiling of amino acid response uncovers unique methionine-deprived response dependent on intact creatine biosynthesis.

    PubMed

    Tang, Xiaohu; Keenan, Melissa M; Wu, Jianli; Lin, Chih-An; Dubois, Laura; Thompson, J Will; Freedland, Stephen J; Murphy, Susan K; Chi, Jen-Tsan

    2015-04-01

    Besides being building blocks for protein synthesis, amino acids serve a wide variety of cellular functions, including acting as metabolic intermediates for ATP generation and for redox homeostasis. Upon amino acid deprivation, free uncharged tRNAs trigger GCN2-ATF4 to mediate the well-characterized transcriptional amino acid response (AAR). However, it is not clear whether the deprivation of different individual amino acids triggers identical or distinct AARs. Here, we characterized the global transcriptional response upon deprivation of one amino acid at a time. With the exception of glycine, which was not required for the proliferation of MCF7 cells, we found that the deprivation of most amino acids triggered a shared transcriptional response that included the activation of ATF4, p53 and TXNIP. However, there was also significant heterogeneity among different individual AARs. The most dramatic transcriptional response was triggered by methionine deprivation, which activated an extensive and unique response in different cell types. We uncovered that the specific methionine-deprived transcriptional response required creatine biosynthesis. This dependency on creatine biosynthesis was caused by the consumption of S-Adenosyl-L-methionine (SAM) during creatine biosynthesis that helps to deplete SAM under methionine deprivation and reduces histone methylations. As such, the simultaneous deprivation of methionine and sources of creatine biosynthesis (either arginine or glycine) abolished the reduction of histone methylation and the methionine-specific transcriptional response. Arginine-derived ornithine was also required for the complete induction of the methionine-deprived specific gene response. Collectively, our data identify a previously unknown set of heterogeneous amino acid responses and reveal a distinct methionine-deprived transcriptional response that results from the crosstalk of arginine, glycine and methionine metabolism via arginine

  10. Arginine and lysine decarboxylases and the acid tolerance response of Salmonella Typhimurium.

    PubMed

    Alvarez-Ordóñez, Avelino; Fernández, Ana; Bernardo, Ana; López, Mercedes

    2010-01-01

    Salmonella Typhimurium CECT 443 inactivation at pH 2.5 in Mineral Medium (MM) and MM supplemented with 0.01% (w/v) arginine, lysine or glutamic acid was studied using stationary-phase cells grown in buffered BHI pH 7.0 (non-acid adapted cells) and acidified BHI up to pH 4.5 with acetic, citric, lactic and hydrochloric acids (acid adapted cells). In all cases, acid adapted cells, with D-values ranging from 23.34 to 86.90 min, showed a significantly higher acid resistance than non-acid adapted cells, with D-values between 8.90 and 10.29 min. Whereas the conditions used for acid adaptation did not exert a significant effect on the acid resistance of the S. Typhimurium CECT 443 resulting cells, the inclusion of lysine and arginine in the challenge medium protected them against acid inactivation, reaching D-values of about 2 and 3 times higher, respectively, than those found in MM or MM supplemented with glutamic acid. None of these three amino acids significantly modified the acid resistance of non-acid adapted cells. The relative expression level of adiA (encoding the arginine decarboxylase), adiY (encoding the transcriptional activator of adiA), cadA (encoding the lysine decarboxylase) and cadB (encoding the lysine/cadaverine transport protein) was examined by quantitative PCR. Acid adapted cells showed higher relative expression levels for both systems, arginine decarboxylase and lysine decarboxylase, which demonstrates that the induction of specialized pH-homeostatic systems plays an important role in S. Typhimurium CECT 443 protection against acid stress. However, the increased acid resistance showed by acid adapted cells challenged in MM arginine or lysine free suggests the existence of other microbial survival strategies.

  11. Available lysine and digestible amino acid contents of proteinaceous foods of India.

    PubMed

    Rutherfurd, Shane M; Bains, Kiran; Moughan, Paul J

    2012-08-01

    Cereals and legumes are staple foods in India and are limiting in lysine and sulphur amino acids, respectively. Available lysine loss, due to Maillard-type reactions that may occur during food preparation, exacerbates the problem of lysine deficiency particularly in cereals. Consequently, determining the contents of digestible essential amino acids, particularly lysine, is important. True ileal digestibilities of most amino acids (including total and reactive lysine) were determined for ten food ingredients and eleven foods commonly consumed in India. Semi-synthetic diets each containing either an ingredient or the prepared food as the sole protein source were formulated to contain 100 g kg(-1) protein (75 g kg(-1) for rice-based diets) and fed to growing rats. Titanium dioxide was included as an indigestible marker. Digesta were collected and the amino acid content (including reactive lysine) of diets and ileal digesta determined. Available (digestible reactive) lysine content ranged from 1·9-15·4 g kg(-1) and 1·8-12·7 g kg(-1) across the ingredients and prepared foods respectively. True ileal amino acid digestibility varied widely both across ingredients and prepared foods for each amino acid (on average 60-92 %) and across amino acids within each ingredient and prepared food (overall digestibility 31-96 %). Amino acid digestibility was low for many of the ingredients and prepared foods and consequently digestibility must be considered when assessing the protein quality of poorer quality foods. Given commonly encountered daily energy intakes for members of the Indian population, it is estimated that lysine is limiting for adults in many Indian diets.

  12. One-step biosynthesis of α-keto-γ-methylthiobutyric acid from L-methionine by an Escherichia coli whole-cell biocatalyst expressing an engineered L-amino acid deaminase from Proteus vulgaris.

    PubMed

    Hossain, Gazi Sakir; Li, Jianghua; Shin, Hyun-dong; Du, Guocheng; Wang, Miao; Liu, Long; Chen, Jian

    2014-01-01

    α-Keto-γ-methylthiobutyric acid (KMTB), a keto derivative of l-methionine, has great potential for use as an alternative to l-methionine in the poultry industry and as an anti-cancer drug. This study developed an environment friendly process for KMTB production from l-methionine by an Escherichia coli whole-cell biocatalyst expressing an engineered l-amino acid deaminase (l-AAD) from Proteus vulgaris. We first overexpressed the P. vulgaris l-AAD in E. coli BL21 (DE3) and further optimized the whole-cell transformation process. The maximal molar conversion ratio of l-methionine to KMTB was 71.2% (mol/mol) under the optimal conditions (70 g/L l-methionine, 20 g/L whole-cell biocatalyst, 5 mM CaCl2, 40°C, 50 mM Tris-HCl [pH 8.0]). Then, error-prone polymerase chain reaction was used to construct P. vulgaris l-AAD mutant libraries. Among approximately 104 mutants, two mutants bearing lysine 104 to arginine and alanine 337 to serine substitutions showed 82.2% and 80.8% molar conversion ratios, respectively. Furthermore, the combination of these mutations enhanced the catalytic activity and molar conversion ratio by 1.3-fold and up to 91.4% with a KMTB concentration of 63.6 g/L. Finally, the effect of immobilization on whole-cell transformation was examined, and the immobilized whole-cell biocatalyst with Ca2+ alginate increased reusability by 41.3% compared to that of free cell production. Compared with the traditional multi-step chemical synthesis, our one-step biocatalytic production of KMTB has an advantage in terms of environmental pollution and thus has great potential for industrial KMTB production.

  13. [Protein utilization in lysine-supplemented barley protein and effectiveness of the limiting amino acid lysine in growing pigs].

    PubMed

    Wecke, C; Gebhardt, G

    1982-04-01

    In 57 N-balance experiments with castrated male pigs (20 ... 65 kg live weight) the influence of graded lysine supplements to crushed barley enriched with energy, minerals and vitamins on nitrogen metabolism and lysine effectiveness was tested. Close correlative relations between lysine concentration and the b-value, the NPU-value, N-balance and N-excretion in urine could be detected. In agreement with the law of minimum a constant lysine effectiveness could be observed within the limiting range. The supplemented synthetic lysine distinguished itself by the same effectiveness as the protein-bound barley lysine. When barley supplemented with lysine is used, an amount of lysine supplement should be chosen from the point of view of nutrition physiology which raises the total lysine content to a maximum level of 6.3 g/16 g N because lysine supplementation exceeding this value without the simultaneous supplementation of limiting threonine remains ineffective.

  14. Role of lysine and acidic amino acid residues on the insecticidal activity of Jackbean urease.

    PubMed

    Real-Guerra, Rafael; Carlini, Célia Regina; Stanisçuaski, Fernanda

    2013-09-01

    Canavalia ensiformis has three isoforms of urease: Jackbean urease (JBU), Jackbean urease II and canatoxin. These isoforms present several biological activities, independent from the enzymatic property, such as entomotoxicity and antifungal properties. The entomotoxic activity is a property of the whole protein, as well as of a 10 kDa peptide released by insect digestive enzymes. Here we have used chemical modification to observe the influence of lysines and acidic residues on JBU enzymatic and insecticidal activities. Chemical modification of lysine residues was performed with dimethylamine-borane complex and formaldehyde, and acidic residues were modified by 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide and ethylenediamine. Derivatized ureases, called JBU-Lys (lysine-modified) and JBU-Ac (acidic residues-modified), were assayed for their biochemical and insecticidal properties. Neither modification altered significantly the kinetic parameters analyzed, indicating that no residue critical for the enzyme activity was affected and that the modifications did not incur in any significant structural alteration. On the other hand, both modifications reduced the toxic activity of the native protein fed to Dysdercus peruvianus. The changes observed in the entomotoxic property of the derivatized proteins reflect alterations in different steps of JBU's toxicity towards insects. JBU-Ac is not susceptible to hydrolysis by insect digestive enzymes, hence impairing the release of toxic peptide(s), while JBU-Lys is processed as the native protein. On the other hand, the antidiuretic effect of JBU on Rhodnius prolixus is altered in JBU-Lys, but not in JBU-Ac. Altogether, these data emphasize the role of lysine and acidic residues on the insecticidal properties of ureases.

  15. Biofortification of rice with the essential amino acid lysine: molecular characterization, nutritional evaluation, and field performance

    PubMed Central

    Yang, Qing-qing; Zhang, Chang-quan; Chan, Man-ling; Zhao, Dong-sheng; Chen, Jin-zhu; Wang, Qing; Li, Qian-feng; Yu, Heng-xiu; Gu, Ming-hong; Sun, Samuel Sai-ming; Liu, Qiao-quan

    2016-01-01

    Rice (Oryza sativa L.), a major staple crop worldwide, has limited levels of the essential amino acid lysine. We previously produced engineered rice with increased lysine content by expressing bacterial aspartate kinase and dihydrodipicolinate synthase and inhibiting rice lysine ketoglutarate reductase/saccharopine dehydrogenase activity. However, the grain quality, field performance, and integration patterns of the transgenes in these lysine-enriched lines remain unclear. In the present study, we selected several elite transgenic lines with endosperm-specific or constitutive regulation of the above key enzymes but lacking the selectable marker gene. All target transgenes were integrated into the intragenic region in the rice genome. Two pyramid transgenic lines (High Free Lysine; HFL1 and HFL2) with free lysine levels in seeds up to 25-fold that of wild type were obtained via a combination of the above two transgenic events. We observed a dramatic increase in total free amino acids and a slight increase in total protein content in both pyramid lines. Moreover, the general physicochemical properties were improved in pyramid transgenic rice, but the starch composition was not affected. Field trials indicated that the growth of HFL transgenic rice was normal, except for a slight difference in plant height and grain colour. Taken together, these findings will be useful for the potential commercialization of high-lysine transgenic rice. PMID:27252467

  16. Biofortification of rice with the essential amino acid lysine: molecular characterization, nutritional evaluation, and field performance.

    PubMed

    Yang, Qing-Qing; Zhang, Chang-Quan; Chan, Man-Ling; Zhao, Dong-Sheng; Chen, Jin-Zhu; Wang, Qing; Li, Qian-Feng; Yu, Heng-Xiu; Gu, Ming-Hong; Sun, Samuel Sai-Ming; Liu, Qiao-Quan

    2016-07-01

    Rice (Oryza sativa L.), a major staple crop worldwide, has limited levels of the essential amino acid lysine. We previously produced engineered rice with increased lysine content by expressing bacterial aspartate kinase and dihydrodipicolinate synthase and inhibiting rice lysine ketoglutarate reductase/saccharopine dehydrogenase activity. However, the grain quality, field performance, and integration patterns of the transgenes in these lysine-enriched lines remain unclear. In the present study, we selected several elite transgenic lines with endosperm-specific or constitutive regulation of the above key enzymes but lacking the selectable marker gene. All target transgenes were integrated into the intragenic region in the rice genome. Two pyramid transgenic lines (High Free Lysine; HFL1 and HFL2) with free lysine levels in seeds up to 25-fold that of wild type were obtained via a combination of the above two transgenic events. We observed a dramatic increase in total free amino acids and a slight increase in total protein content in both pyramid lines. Moreover, the general physicochemical properties were improved in pyramid transgenic rice, but the starch composition was not affected. Field trials indicated that the growth of HFL transgenic rice was normal, except for a slight difference in plant height and grain colour. Taken together, these findings will be useful for the potential commercialization of high-lysine transgenic rice.

  17. Development of artificial neural network models based on experimental data of response surface methodology to establish the nutritional requirements of digestible lysine, methionine, and threonine in broiler chicks.

    PubMed

    Mehri, M

    2012-12-01

    An artificial neural network (ANN) approach was used to develop feed-forward multilayer perceptron models to estimate the nutritional requirements of digestible lysine (dLys), methionine (dMet), and threonine (dThr) in broiler chicks. Sixty data lines representing response of the broiler chicks during 3 to 16 d of age to dietary levels of dLys (0.88-1.32%), dMet (0.42-0.58%), and dThr (0.53-0.87%) were obtained from literature and used to train the networks. The prediction values of ANN were compared with those of response surface methodology to evaluate the fitness of these 2 methods. The models were tested using R(2), mean absolute deviation, mean absolute percentage error, and absolute average deviation. The random search algorithm was used to optimize the developed ANN models to estimate the optimal values of dietary dLys, dMet, and dThr. The ANN models were used to assess the relative importance of each dietary input on the bird performance using sensitivity analysis. The statistical evaluations revealed the higher accuracy of ANN to predict the bird performance compared with response surface methodology models. The optimization results showed that the maximum BW gain may be obtained with dietary levels of 1.11, 0.51, and 0.78% of dLys, dMet, and dThr, respectively. Minimum feed conversion ratio may be achieved with dietary levels of 1.13, 0.54, 0.78% of dLys, dMet, and dThr, respectively. The sensitivity analysis on the models indicated that dietary Lys is the most important variable in the growth performance of the broiler chicks, followed by dietary Thr and Met. The results of this research revealed that the experimental data of a response-surface-methodology design could be successfully used to develop the well-designed ANN for pattern recognition of bird growth and optimization of nutritional requirements. The comparison between the 2 methods also showed that the statistical methods may have little effect on the ideal ratios of dMet and dThr to dLys in

  18. Methionine and Choline Supply during the Periparturient Period Alter Plasma Amino Acid and One-Carbon Metabolism Profiles to Various Extents: Potential Role in Hepatic Metabolism and Antioxidant Status

    PubMed Central

    Zhou, Zheng; Vailati-Riboni, Mario; Luchini, Daniel N.; Loor, Juan J.

    2016-01-01

    The objective of this study was to profile plasma amino acids (AA) and derivatives of their metabolism during the periparturient period in response to supplemental rumen-protected methionine (MET) or rumen-protected choline (CHOL). Forty cows were fed from −21 through 30 days around parturition in a 2 × 2 factorial design a diet containing MET or CHOL. MET supply led to greater circulating methionine and proportion of methionine in the essential AA pool, total AA, and total sulfur-containing compounds. Lysine in total AA also was greater in these cows, indicating a better overall AA profile. Sulfur-containing compounds (cystathionine, cystine, homocystine, and taurine) were greater in MET-fed cows, indicating an enriched sulfur-containing compound pool due to enhanced transsulfuration activity. Circulating essential AA and total AA concentrations were greater in cows supplied MET due to greater lysine, arginine, tryptophan, threonine, proline, asparagine, alanine, and citrulline. In contrast, CHOL supply had no effect on essential AA or total AA, and only tryptophan and cystine were greater. Plasma 3-methylhistidine concentration was lower in response to CHOL supply, suggesting less tissue protein mobilization in these cows. Overall, the data revealed that enhanced periparturient supply of MET has positive effects on plasma AA profiles and overall antioxidant status. PMID:28036059

  19. Methionine Biosynthesis in Lemna

    PubMed Central

    Thompson, Gregory A.; Datko, Anne H.; Mudd, S. Harvey; Giovanelli, John

    1982-01-01

    Regulation of enzymes of methionine biosynthesis was investigated by measuring the specific activities of O-phosphohomoserine-dependent cystathionine γ-synthase, O-phosphohomoserine sulfhydrylase, and O-acetylserine sulfhydrylase in Lemna paucicostata Hegelm. 6746 grown under various conditions. For cystathionine γ-synthase, it was observed that (a) adding external methionine (2 μm) decreased specific activity to 15% of control, (b) blocking methionine synthesis with 0.05 μml-aminoethoxyvinylglycine or with 36 μm lysine plus 4 μm threonine (Datko, Mudd 1981 Plant Physiol 69: 1070-1076) caused a 2- to 3-fold increase in specific activity, and (c) blocking methionine synthesis and adding external methionine led to the decreased specific activity characteristic of methionine addition alone. Activity in extracts from control cultures was unaffected by addition of methionine, lysine, threonine, lysine plus threonine, S-adenosylmethionine, or S-methylmethionine sulfonium to the assay mixture. Parallel studies of O-phosphohomoserine sulfhydrylase and O-acetylserine sulfhydrylase showed that O-phosphohomoserine sulfhydrylase activity responded to growth conditions identically to cystathionine γ-synthase activity, whereas O-acetylserine sulfhydrylase activity remained unaffected. Lemna extracts did not catalyze lanthionine formation from O-acetylserine and cysteine. Estimates of kinetic constants for the three enzyme activities indicate that O-acetylserine sulfhydrylase has much higher activity and affinity for sulfide than O-phosphohomoserine sulfhydrylase. The results suggest that (a) methionine, or one of its products, regulates the amount of active cystathionine γ-synthase in Lemna, (b) O-phosphohomoserine sulfhydrylase and cystathionine γ-synthase are probably activities of one enzyme that has low specificity for its sulfur-containing substrate, and (c) O-acetylserine sulfhydrylase is a separate enzyme. The relatively high activity and affinity for sulfide of

  20. YjeH Is a Novel Exporter of l-Methionine and Branched-Chain Amino Acids in Escherichia coli.

    PubMed

    Liu, Qian; Liang, Yong; Zhang, Yun; Shang, Xiuling; Liu, Shuwen; Wen, Jifu; Wen, Tingyi

    2015-11-01

    Amino acid efflux transport systems have important physiological functions and play vital roles in the fermentative production of amino acids. However, no methionine exporter has yet been identified in Escherichia coli. In this study, we identified a novel amino acid exporter, YjeH, in E. coli. The yjeH overexpression strain exhibited high tolerance to the structural analogues of l-methionine and branched-chain amino acids, decreased intracellular amino acid levels, and enhanced export rates in the presence of a Met-Met, Leu-Leu, Ile-Ile, or Val-Val dipeptide, suggesting that YjeH functions as an exporter of l-methionine and the three branched-chain amino acids. The export of the four amino acids in the yjeH overexpression strain was competitively inhibited in relation to each other. The expression of yjeH was strongly induced by increasing cytoplasmic concentrations of substrate amino acids. Green fluorescent protein (GFP)-tagged YjeH was visualized by total internal reflection fluorescence microscopy to confirm the plasma membrane localization of YjeH. Phylogenetic analysis of transporters indicated that YjeH belongs to the amino acid efflux family of the amino acid/polyamine/organocation (APC) superfamily. Structural modeling revealed that YjeH has the typical "5 + 5" transmembrane α-helical segment (TMS) inverted-repeat fold of APC superfamily transporters, and its binding sites are strictly conserved. The enhanced capacity of l-methionine export by the overexpression of yjeH in an l-methionine-producing strain resulted in a 70% improvement in titer. This study supplements the transporter classification and provides a substantial basis for the application of the methionine exporter in metabolic engineering.

  1. YjeH Is a Novel Exporter of l-Methionine and Branched-Chain Amino Acids in Escherichia coli

    PubMed Central

    Liu, Qian; Liang, Yong; Zhang, Yun; Shang, Xiuling; Liu, Shuwen; Wen, Jifu

    2015-01-01

    Amino acid efflux transport systems have important physiological functions and play vital roles in the fermentative production of amino acids. However, no methionine exporter has yet been identified in Escherichia coli. In this study, we identified a novel amino acid exporter, YjeH, in E. coli. The yjeH overexpression strain exhibited high tolerance to the structural analogues of l-methionine and branched-chain amino acids, decreased intracellular amino acid levels, and enhanced export rates in the presence of a Met-Met, Leu-Leu, Ile-Ile, or Val-Val dipeptide, suggesting that YjeH functions as an exporter of l-methionine and the three branched-chain amino acids. The export of the four amino acids in the yjeH overexpression strain was competitively inhibited in relation to each other. The expression of yjeH was strongly induced by increasing cytoplasmic concentrations of substrate amino acids. Green fluorescent protein (GFP)-tagged YjeH was visualized by total internal reflection fluorescence microscopy to confirm the plasma membrane localization of YjeH. Phylogenetic analysis of transporters indicated that YjeH belongs to the amino acid efflux family of the amino acid/polyamine/organocation (APC) superfamily. Structural modeling revealed that YjeH has the typical “5 + 5” transmembrane α-helical segment (TMS) inverted-repeat fold of APC superfamily transporters, and its binding sites are strictly conserved. The enhanced capacity of l-methionine export by the overexpression of yjeH in an l-methionine-producing strain resulted in a 70% improvement in titer. This study supplements the transporter classification and provides a substantial basis for the application of the methionine exporter in metabolic engineering. PMID:26319875

  2. Characterization and detection of lysine-arginine cross-links derived from dehydroascorbic acid.

    PubMed

    Reihl, Oliver; Lederer, Markus O; Schwack, Wolfgang

    2004-02-25

    Covalently cross-linked proteins are among the major modifications caused by the advanced Maillard reaction. So far, the chemical nature of these aggregates is largely unknown. L-dehydroascorbic acid (DHA, 5), the oxidation product of L-ascorbic acid (vitamin C), is known as a potent glycation agent. Identification is reported for the lysine-arginine cross-links N6-[2-[(4-amino-4-carboxybutyl)amino]-5-(2-hydroxyethyl)-3,5-dihydro-4H-imidazol-4-ylidene]-L-lysine (9), N6-[2-[(4-amino-4-carboxybutyl)amino]-5-(1,2-dihydroxyethyl)-3,5-dihydro-4H-imidazol-4-ylidene]-L-lysine (11), and N6-[2-[(4-amino-4-carboxybutyl)amino]-5-[(1S,2S)-1,2,3-trihydroxypropyl]-3,5-dihydro-4H-imidazol-4-ylidene]-L-lysine (13). The formation pathways could be established starting from dehydroascorbic acid (5), the degradation products 1,3,4-trihydroxybutan-2-one (7, L-erythrulose), 3,4-dihydroxy-2-oxobutanal (10, L-threosone), and L-threo-pentos-2-ulose (12, L-xylosone) were proven as precursors of the lysine-arginine cross-links 9, 11, and 13. Products 9 and 11 were synthesized starting from DHA 5, compound N6-[2-[(4-amino-4-carboxybutyl)amino]-5-[(1S,2R)-1,2,3-trihydroxypropyl]-3,5-dihydro-4H-imidazol-4-ylidene]-L-lysine (16) via the precursor D-erythro-pentos-2-ulose (15). The present study revealed that the modification of lysine and arginine side chains by DHA 5 is a complex process and could involve a number of reactive carbonyl species.

  3. Modulation of benzodiazepine by lysine and pipecolic acid on pentylenetetrazol-induced seizures

    SciTech Connect

    Chang, Y.F.; Hargest, V.; Chen, J.S.

    1988-01-01

    L-lysine and its metabolite pipecolic acid (PA) have been studied for their effects on pentylenetetrazol (PTZ)-induced seizures in mice. L-Lysine of L-Pa i.p. significantly increased clonic and tonic latencies in a dose-dependent manner against 90 mg/kg PTZ-induced seizures. L-Lysine but not L-Pa enhanced the anticonvulsant effect of diazepam (DZ). L-Pa i.c.v. showed a slight decrease in clonic latency; it did not enhance the antiseizure activity of DZ; it caused seizures at 0.6 mmol/kg. D-PA i.c.v. displayed an opposite effect compared to its L-isomer. The anticonvulsant effect of L-lysine in terms of increase in seizure latency and survival was even more amplified when tested with a submaximal PTZ concentration. L-Lysine showed an enhancement of specific /sup 3/H-flunitrazepam(FZ) binding to mouse brain membranes both in vitro an din vivo. The possibility of L-lysine acting as a modulator for the GABA/benzodiazepine receptors was demonstrated. Since L-PA showed enhancement of /sup 3/H-FZ binding only in vitro but not in vivo, the anticonvulsant effect of L-PA may not be linked to the GABA/benzodiazepine receptor.

  4. Effects of calcium soaps of rapeseed fatty acids and protected methionine on milk yield and composition in dairy cows.

    PubMed

    Kowalski, Z M; Pisulewski, P M; Spanghero, M

    1999-11-01

    The objective of this study was to determine the effects of supplementing the diets of dairy cows with Ca soaps of rapeseed fatty acids (CSRFA) and rumen-protected (RP) methionine on their milk yield and composition, including milk protein fractions and fatty acids. Twelve Polish Red Lowland cows were used in a complete balanced two period changeover experiment. The four treatment diets were a control consisting of a total mixed ration of grass silage and concentrates, and the total mixed ration supplemented with RP methionine, CSRFA or RP methionine plus CSRFA. Dry matter intake was not affected by diet. Milk yield increased when cows were given the diet with CSRFA, but supplementation of diets with RP methionine did not affect milk yield. Milk protein content, but not milk protein yield, decreased when CSRFA was given. The addition of RP methionine to the control diet and the CSRFA diet produced similar increases in the milk protein content. Supplementation of the diet with CSRFA significantly changed the milk fatty acid profile: the proportions of 10:0, 12:0, 14:0, 15:0 and 16:0 in milk fat decreased, but those of 18:0 and cis-18:1 increased. We conclude that CSRFA can be used in practical dairy diets to increase milk yield and manipulate its fatty acid composition.

  5. The in vivo sparing of methionine by cysteine in sulfur amino acid requirements in animal models and adult humans.

    PubMed

    Ball, Ronald O; Courtney-Martin, Glenda; Pencharz, Paul B

    2006-06-01

    Sulfur amino acid metabolism has been receiving increased attention because of the link to chronic diseases such as cardiovascular disease, Alzheimer's disease, and diabetes. In addition, the role of cysteine and optimal intakes for physiological substrates such as glutathione are currently of considerable interest in human health. Although the dietary indispensability of methionine is not in question, the ability of cysteine to substitute for a portion of its requirement has been the topic of much debate. Methionine is often the most limiting amino acid in the diets of the developing world's population because of its low concentration in cereal grains. Therefore, the ability of cysteine to substitute for methionine requirement is not just biologically interesting; it is also of considerable economic and social importance. The primary goal of this review is to discuss the available evidence on the effect of cysteine substitution for methionine to meet the total sulfur amino acid requirement in adult humans, including an assessment of the methodological features of experiments with conflicting results. Assessment of the requirement experiments for amino acids with complex metabolism such as methionine and cysteine must begin with a careful definition of requirements and what substitution means. As a result of these definitions, a set of criteria for the intakes of methionine that will allow demonstration of the substitution effect have been developed. Some recent publications are assessed using these definitions and criteria, and a possible reason for the conflicting results in the literature is proposed. An approach to estimating tolerable upper intakes is also proposed. Research on in vivo sulfur amino acid metabolism in humans is tremendously difficult, and therefore, we do not wish to be overly critical of the high-quality work of the ambitious and highly intelligent men and women who have conducted various studies. Our goal is to objectively review the data for

  6. SucStruct: Prediction of succinylated lysine residues by using structural properties of amino acids.

    PubMed

    López, Yosvany; Dehzangi, Abdollah; Lal, Sunil Pranit; Taherzadeh, Ghazaleh; Michaelson, Jacob; Sattar, Abdul; Tsunoda, Tatsuhiko; Sharma, Alok

    2017-03-28

    Post-Translational Modification (PTM) is a biological reaction which contributes to diversify the proteome. Despite many modifications with important roles in the cellular activity, lysine succinylation has recently emerged as an important PTM mark. It alters the chemical structure of lysines, leading to remarkable changes in the structure and function of proteins. Given the huge amount of proteins being sequenced in the post-genome era, the experimental detection of succinylated residues remains expensive, inefficient and time-consuming. Therefore, the development of computational tools for accurately predicting succinylated lysines is an urgent necessity. To date, several approaches have been proposed but their sensitivity has been reportedly poor. In this paper, we propose an approach that utilizes structural features of amino acids to improve lysine succinylation prediction. Succinylated and non-succinylated lysines were first retrieved from 670 proteins and characteristics such as accessible surface area, backbone torsion angles, and local structure conformations were incorporated. We used the k-nearest neighbors cleaning for dealing with class imbalance and designed a pruned decision tree for classification. Our predictor, referred as SucStruct (Succinylation using Structural features), proved to significantly improve performance when compared to previous predictors, with sensitivity, accuracy and Mathew's correlation coefficient equal to 0.7334-0.7946, 0.7444-0.7608 and 0.4884-0.5240, respectively.

  7. In vitro degradation of lysine by ruminal fluid-based fermentations and by Fusobacterium necrophorum.

    PubMed

    Elwakeel, E A; Amachawadi, R G; Nour, A M; Nasser, M E A; Nagaraja, T G; Titgemeyer, E C

    2013-01-01

    The objective of these studies was to characterize some factors affecting lysine degradation by mixed ruminal bacteria and by ruminal Fusobacterium necrophorum. Mixed ruminal bacteria degraded lysine, and addition of pure cultures of F. necrophorum did not increase lysine degradation. Addition of acetic or propionic acid strikingly reduced NH(3) production from lysine by mixed ruminal bacteria at pH 6, but not at pH 7. Although typical ruminal environments with acidic pH and normal concentrations of volatile fatty acids might inhibit lysine degradation by F. necrophorum, ruminal fluid contained enough bacteria with a lysine-degrading capacity to ferment 50 mM lysine in vitro. Of 7 strains of ruminal F. necrophorum tested, all grew on both lactate and lysine as the primary energy source. Both subspecies of ruminal F. necrophorum (necrophorum and funduliforme) used lysine as a primary C and energy source. Lysine and glutamic acid were effectively fermented by F. necrophorum, but alanine and tryptophan were not, and histidine and methionine were fermented only to a minor extent. The end products of lactate fermentation by F. necrophorum were propionate and acetate, and those of lysine degradation were butyrate and acetate. Fermentation of glutamic acid by F. necrophorum yielded acetate and butyrate in a ratio near to 2:1. The minimum inhibitory concentration of tylosin for F. necrophorum was not dependent on whether bacteria were grown with lactate or lysine, but F. necrophorum was more susceptible to monensin when grown on lysine than on lactate. Although F. necrophorum is generally resistant to monensin, the ionophore may reduce lysine degradation by F. necrophorum in the rumen. The essential oil components limonene, at 20 or 100 μg/mL, and thymol, at 100 μg/mL, inhibited F. necrophorum growth, whereas eugenol, guaiacol, and vanillin had no effect. Our findings may lead to ways to minimize ruminal lysine degradation and thus increase its availability to the animal.

  8. Pyridoxal 5'-phosphate inactivates DNA topoisomerase IB by modifying the lysine general acid.

    PubMed

    Vermeersch, Jacqueline J; Christmann-Franck, Serge; Karabashyan, Leon V; Fermandjian, Serge; Mirambeau, Gilles; Der Garabedian, P Arsène

    2004-01-01

    The present results demonstrate that pyridoxal, pyridoxal 5'-phosphate (PLP) and pyridoxal 5'-diphospho-5'-adenosine (PLP-AMP) inhibit Candida guilliermondii and human DNA topoisomerases I in forming an aldimine with the epsilon-amino group of an active site lysine. PLP acts as a competitive inhibitor of C.guilliermondii topoisomerase I (K(i) = 40 microM) that blocks the cleavable complex formation. Chemical reduction of PLP-treated enzyme reveals incorporation of 1 mol of PLP per mol of protein. The limited trypsic proteolysis releases a 17 residue peptide bearing a lysine-bound PLP (KPPNTVIFDFLGK*DSIR). Targeted lysine (K*) in C.guilliermondii topoisomerase I corresponds to that found in topoisomerase I of Homo sapiens (K532), Candida albicans (K468), Saccharomyces cerevisiae (K458) and Schizosaccharomyces pombe (K505). In the human enzyme, K532, belonging to the active site acts as a general acid catalyst and is therefore essential for activity. The spatial orientation of K532-PLP within the active site was approached by molecular modeling using available crystallographic data. The PLP moiety was found at close proximity of several active residues. PLP could be involved in the cellular control of topoisomerases IB. It constitutes an efficient tool to explore topoisomerase IB dynamics during catalysis and is also a lead for new drugs that trap the lysine general acid.

  9. A Single Amino Acid Change Is Responsible for Evolution of Acyltransferase Specificity in Bacterial Methionine Biosynthesis

    SciTech Connect

    Zubieta, C.; Arkus, K.A.J.; Cahoon, R.E.; Jez, J.M.

    2009-05-28

    Bacteria and yeast rely on either homoserine transsuccinylase (HTS, metA) or homoserine transacetylase (HTA; met2) for the biosynthesis of methionine. Although HTS and HTA catalyze similar chemical reactions, these proteins are typically unrelated in both sequence and three-dimensional structure. Here we present the 2.0 {angstrom} resolution x-ray crystal structure of the Bacillus cereus metA protein in complex with homoserine, which provides the first view of a ligand bound to either HTA or HTS. Surprisingly, functional analysis of the B. cereus metA protein shows that it does not use succinyl-CoA as a substrate. Instead, the protein catalyzes the transacetylation of homoserine using acetyl-CoA. Therefore, the B. cereus metA protein functions as an HTA despite greater than 50% sequence identity with bona fide HTS proteins. This result emphasizes the need for functional confirmation of annotations of enzyme function based on either sequence or structural comparisons. Kinetic analysis of site-directed mutants reveals that the B. cereus metA protein and the E. coli HTS share a common catalytic mechanism. Structural and functional examination of the B. cereus metA protein reveals that a single amino acid in the active site determines acetyl-CoA (Glu-111) versus succinyl-CoA (Gly-111) specificity in the metA-like of acyltransferases. Switching of this residue provides a mechanism for evolving substrate specificity in bacterial methionine biosynthesis. Within this enzyme family, HTS and HTA activity likely arises from divergent evolution in a common structural scaffold with conserved catalytic machinery and homoserine binding sites.

  10. Biomimetic niche for neural stem cell differentiation using poly-L-lysine/hyaluronic acid multilayer films.

    PubMed

    Lee, I-Chi; Wu, Yu-Chieh; Cheng, En-Ming; Yang, Wen-Ting

    2015-05-01

    Polyelectrolyte multilayer films have been suggested as tunable substrates with flexible surface properties that can modulate cell behavior. However, these films' biological effects on neural stem/progenitor cells have rarely been studied. Herein, biomimetic multilayer films composed of hyaluronic acid and poly-L-lysine were chosen to mimic the native extracellular matrix niche of brain tissue and were evaluated for their inductive effects, without the addition of chemical factors. Because neural stem/progenitor cells are sensitive to substrate properties, it is important that this system provides control over the surface charge, and slight stiffness variations are also possible. Both of these factors affect neural stem/progenitor cell differentiation. The results showed that neural stem/progenitor cells were induced to differentiate on the poly-L-lysine/hyaluronic acid multilayer films with 0.5-4 alternating layers. In addition, the neurite outgrowth length was regulated by the surface charge of the terminal layer but did not increase with the layer number. In contrast, the quantity of differentiated neurons was enhanced slightly as the number of layers increased but was not affected by the surface charge of the terminal layer. In sum, material pairs in the form of native poly-L-lysine/hyaluronic acid films achieved important targets for neural regenerative medicine, including enhancement of the neurite outgrowth length, regulation of neuron differentiation, and the formation of a network. These extracellular matrix-mimetic poly-L-lysine/hyaluronic acid multilayer films may provide a versatile platform that could be useful for surface modification for applications in neural engineering.

  11. [Nutrient composition of some newly bred high-protein and/or high-lysine cereal strains and digestibility by growing pigs. 2. Protein quality and digestibility of the amino acids].

    PubMed

    Bock, H D; Wünsche, J; Meinl, M; Hennig, U; Völker, T

    1982-05-01

    After a first report on the nutrient composition and the digestibility of some newly bred varieties of cereals, information on the apparent digestibility of the amino acids in growing female pigs ascertained with the same test material is given. These data serve the evaluation of the quality of such cultivation products and the completion of the data in of feedstuffs tables and tables of the amino acid content. The newly bred barleystrains meet the requirement of digestible protein of pigs, wheat exceeds it whereas the maize samples fall short of it. The content of digestible lysine in the newly bred barleystrain approximate the requirement of growing pigs. There is ample digestible methionine + cystine in all samples. This again proves that newly cultivated barley and wheat rich in protein and lysine if accordingly supplemented with the limiting amino acid lysine can be used as sole feed for fattening pigs. Moreover, it becomes again obvious that the apparent digestibility of the amino acids can, for practical purposes, be sufficiently exactly evaluated with the help of the apparent digestibility of the crude protein of the respective feedstuff.

  12. Reversible lysine modification on proteins by using functionalized boronic acids.

    PubMed

    Cal, Pedro M S D; Frade, Raquel F M; Cordeiro, Carlos; Gois, Pedro M P

    2015-05-26

    Iminoboronates have been utilized to successfully install azide and alkyne bioorthogonal functions on proteins, which may then be further reacted with their bioorthogonal counterparts. These constructs were also used to add polyethylene glycol (PEG) to insulin, a modification which has been shown to be reversible in the presence of fructose. Finally, iminoboronates were used to assemble a folic acid/paclitaxel small-molecule/drug conjugate in situ with an IC50  value of 20.7 nM against NCI-H460 cancer cells and negligible cytotoxicity against the CRL-1502 noncancer cells.

  13. Chiral effects in amino acid adsorption on Au(111): A comparison of cysteine, homocysteine and methionine

    NASA Astrophysics Data System (ADS)

    Popa, Tatiana; Ting, Elvis C. M.; Paci, Irina

    2014-11-01

    A combined classical/quantum methodology is used to examine chiral effects upon adsorption of three sulfur-containing amino acids on the Au(111) surface: cysteine, homocysteine and methionine. Parallel tempering Monte Carlo simulations were employed to broadly examine the configurational space of monomers, dimers and trimers of the molecules on the gold surface. Density functional theory was applied to promising structural targets in order to incorporate higher order electronic structure effects in a study of relative stabilities of the various molecular states upon adsorption. As the precursors of chiral structure formation, like and unlike dimers were investigated at some length, with consideration given to the mode of sorption (chemisorption of physisorption) and the existence of zwitterionic states. We found that neutral (non-zwitterionic) molecules adsorbed weakly on the highly-coordinated Au(111) surfaces. As a consequence, pair configurations in dimers were insufficiently constrained to lead to differential stabilities of homochiral and heterochiral dimers. Whereas neutral molecule interactions were non-discriminating, strong chiral discrimination was found in zwitterionic amino acids. The zwitterionic forms of the larger molecules equilibrated closer to the surface, and the stronger molecule-molecule and molecule-surface interactions were such that homochiral dimers were stable whereas heterochiral dimers were not.

  14. Effects of Supplemental Liquid DL-methionine Hydroxy Analog Free Acid in Diet on Growth Performance and Gastrointestinal Functions of Piglets

    PubMed Central

    Kaewtapee, C.; Krutthai, N.; Bunchasak, C.

    2016-01-01

    This study was conducted to determine the effect of dietary supplementation of liquid DL-methionine hydroxy analog free acid (DL-MHA) on growth performance and gastrointestinal conditions of piglets. One hundred and eighty crossbred barrow piglets (Large White×Landrace, body weight: 12.48±0.33 kg) were divided into three groups with ten replications of six piglets each. Piglets received DL-MHA in diet at a concentration of 0 (control group), 0.15%, or 0.24%. The results indicated that increasing the standardized ileal digestible (SID) of sulfur amino acids (SAA) to lysine (SID SAA:Lys) ratio by supplementation of DL-MHA tended to increase (quadratic; p<0.10) weight gain and ADG, and showed slightly greater (linear; p<0.10) gain:feed ratio. The pH in the diet and cecum linearly decreased (p<0.01), whereas pH in colon had a quadratic response (p<0.01) with increasing supplementation of DL-MHA. By greater supplementation of DL-MHA, the population of Lactobacillus spp. in rectum was likely to increase (quadratic; p<0.10), but Escherichia coli population in the diet was reduced (quadratic; p<0.05). Acetic acid concentration and total short-chain fatty acids in cecum linearly increased (p<0.05), whereas valeric acid in cecum quadratically increased (p<0.05) with increasing DL-MHA levels. Moreover, the villous height of the jejunum quadratically increased (p<0.01) as the supplementation of DL-MHA was increased. It is concluded that the addition of DL-MHA in diet improved the growth performance and the morphology of gastrointestinal tract of piglets. PMID:26954213

  15. Photooxidation of Methionine

    ERIC Educational Resources Information Center

    Lewis, Catherine; Scouten, William H.

    1976-01-01

    Describes an experiment in which the photooxidation of methionine using free methylene blue as the sensitizer is applied to the isolated amino acid or to the methionyl residues of a complex polypeptide. (MLH)

  16. Mesoporous silica nanoparticles functionalized with folic acid/methionine for active targeted delivery of docetaxel

    PubMed Central

    Khosravian, Pegah; Shafiee Ardestani, Mehdi; Khoobi, Mehdi; Ostad, Seyed Naser; Dorkoosh, Farid Abedin; Akbari Javar, Hamid; Amanlou, Massoud

    2016-01-01

    Mesoporous silica nanoparticles (MSNs) are known as carriers with high loading capacity and large functionalizable surface area for target-directed delivery. In this study, a series of docetaxel-loaded folic acid- or methionine-functionalized mesoporous silica nanoparticles (DTX/MSN-FA or DTX/MSN-Met) with large pores and amine groups at inner pore surface properties were prepared. The results showed that the MSNs were successfully synthesized, having good pay load and pH-sensitive drug release kinetics. The cellular investigation on MCF-7 cells showed better performance of cytotoxicity and cell apoptosis and an increase in cellular uptake of targeted nanoparticles. In vivo fluorescent imaging on healthy BALB/c mice proved that bare MSN-NH2 are mostly accumulated in the liver but MSN-FA or MSN-Met are more concentrated in the kidney. Importantly, ex vivo fluorescent images of tumor-induced BALB/c mice organs revealed the ability of MSN-FA to reach the tumor tissues. In conclusion, DTX/MSNs exhibited a good anticancer activity and enhanced the possibility of targeted drug delivery for breast cancer. PMID:27980423

  17. Lysine metabolism in antisense C-hordein barley grains.

    PubMed

    Schmidt, Daiana; Rizzi, Vanessa; Gaziola, Salete A; Medici, Leonardo O; Vincze, Eva; Kozak, Marcin; Lea, Peter J; Azevedo, Ricardo A

    2015-02-01

    The grain proteins of barley are deficient in lysine and threonine due to their low concentrations in the major storage protein class, the hordeins, especially in the C-hordein subgroup. Previously produced antisense C-hordein transgenic barley lines have an improved amino acid composition, with increased lysine, methionine and threonine contents. The objective of the study was to investigate the possible changes in the regulation of key enzymes of the aspartate metabolic pathway and the contents of aspartate-derived amino acids in the nontransgenic line (Hordeum vulgare L. cv. Golden Promise) and five antisense C-hordein transgenic barley lines. Considering the amounts of soluble and protein-bound aspartate-derived amino acids together with the analysis of key enzymes of aspartate metabolic pathway, we suggest that the C-hordein suppression did not only alter the metabolism of at least one aspartate-derived amino acid (threonine), but major changes were also detected in the metabolism of lysine and methionine. Modifications in the activities and regulation of aspartate kinase, dihydrodipicolinate synthase and homoserine dehydrogenase were observed in most transgenic lines. Furthermore the activities of lysine α-ketoglutarate reductase and saccharopine dehydrogenase were also altered, although the extent varied among the transgenic lines.

  18. Effects of reducing dietary protein, methionine, choline, folic acid, and vitamin B12 during the late stages of the egg production cycle on performance and eggshell quality.

    PubMed

    Keshavarz, K

    2003-09-01

    A series of four experiments was conducted to determine whether-shell quality during the late stages of egg production can be improved by using diets that are effective in reducing egg size. The experiments involved dietary manipulation of protein, methionine, choline, folic acid, and vitamin B12. In experiment 1, reducing dietary protein in combination of reducing the dietary methionine and choline or this diet without supplemental folic acid and vitamin B12 resulted in reduced egg weight and improved shell quality. However, egg production also was drastically reduced. In experiment 2, reducing the dietary level of methionine, without adding supplemental choline, folic acid, and vitamin B12 reduced egg size and improved shell quality, but egg production was reduced as well. In this experiment reducing the dietary methionine without supplemental folic acid and vitamin B12 reduced egg size and improved shell quality with no adverse effect on egg production. In experiment 3, reducing the dietary level of methionine and choline or reducing the dietary level of choline, folic acid, and vitamin B12 reduced egg size and improved shell quality without adverse effects on egg production. On the other hand, reducing dietary methionine, folic acid, vitamin B12, and supplemental choline reduced egg weight and improved shell quality but lowered egg production. In experiment 4, reducing dietary methionine together with reducing choline and vitamin B12 reduced egg size and improved shell quality with no adverse effect on egg production. The results of this series of experiments generally indicate that certain manipulations of the combination of methionine, choline, folic acid, and vitamin B12 have the potential to reduce egg weight and improve shell quality without affecting egg production during the latter stages of the egg production cycle.

  19. Contribution of gut microbial lysine to liver and milk amino acids in lactating does.

    PubMed

    Abecia, Leticia; Balcells, Joaquím; Fondevila, Manuel; Belenguer, Alvaro; Holtrop, Grietje; Lobley, Gerald E

    2008-11-01

    The contribution of microbial amino acids through caecotrophy to tissue protein metabolism was investigated in lactating does. Attempts were made to vary microbial supply through a dietary antibiotic, Zn bacitracin, and to vary tissue demand through manipulation of litter size. Three groups of eight New Zealand does were fed different experimental diets from day 28 of pregnancy to day 26 of lactation. The control group received the basal diet formulated to meet requirements with grass hay, wheat, soybean meal and barley grain. The second (no antibiotic) group and the third (bacitracin; BAC) group ingested the basal diet supplemented with ammonium sulfate (5 g/kg), initially unlabelled (day 1 to day 8) then labelled with 15N (day 9 to day 30), while the BAC diet was also supplemented throughout with antibiotic (Zn bacitracin; 100 mg/kg). From just after birth each group of does was subdivided into two groups, each of four females, with the litter size either five (LS5) or nine (LS9) pups. The 15N enrichment in liver, milk and caecal bacteria amino acids was determined by GC-combustion-isotope ratio MS. All amino acids in bacterial protein were enriched with the (15 NH 4)2SO4 treatment, with lysine 15N enrichment significantly greater in caecal bacteria (0.23 (SE 0.0063) atom % excess (ape)) than in liver (0.04 (SE 0.0004) ape) or milk protein (0.05 (SE 0.0018) ape), confirming the double origin (bacterial and dietary) of tissue lysine. The contribution of microbes to tissue lysine was 0.23 (SE 0.006) when milk protein was used as reference.

  20. Isolation and characterization of a new advanced glycation endproduct of dehydroascorbic acid and lysine.

    PubMed

    Argirov, Ognyan K; Lin, Bin; Olesen, Paul; Ortwerth, Beryl J

    2003-03-17

    Proteins are subject of posttranslational modification by sugars and their degradation products in vivo. The process is often referred as glycation. L-Dehydroascorbic acid (DHA), an oxidation product of L-ascorbic acid (vitamin C), is known as a potent glycation agent. A new product of modification of lysine epsilon -amino group by DHA was discovered as a result of the interaction between Boc-Lys and dehydroascorbic acid. The chromatographic and spectral analyses revealed that the structure of the product was 1-(5-ammonio-5-carboxypentyl)-3-oxido-4-(hydroxymethyl)pyridinium. The same compound was isolated from DHA modified calf lens protein after hydrolysis and chromatographic separation. The study confirmed that L-erythrulose is an important intermediate of modification of proteins by DHA. The structure of the reported product and in vitro experiments suggested that L-erythrulose could further transform to L-threose, L-erythrose and glycolaldehyde under conditions similar to physiological. The present study revealed that the modification of epsilon -amino groups of lysine residues by DHA is a complex process and could involve a number of reactive carbonyl species.

  1. Lysine requirements of pre-lay broiler breeder pullets: determination by indicator amino acid oxidation.

    PubMed

    Coleman, Russell A; Bertolo, Robert F; Moehn, Soenke; Leslie, Michael A; Ball, Ronald O; Korver, Doug R

    2003-09-01

    The indicator amino acid oxidation (IAAO) method allows the determination of amino acid requirements under conditions of low growth rate as found in pre-laying broiler breeder pullets. Cobb 500 breeder pullets (20 wk old; 2290 +/- 280 g, n = 4) were adapted (6 d) to a pelleted, purified control diet containing all nutrients at >or=110% of NRC recommendations. After recovery from surgery for implantation of a jugular catheter, each bird was fed, in random order, test diets containing one of nine levels of lysine (0.48, 0.96, 1.92, 2.88, 3.84, 4.80, 7.68, 9.60 and 14.40 g/kg of diet). Indicator oxidation was determined during 4-h primed (74 kBq/kg body), constant infusions (44 kBq x h(-1). kg body(-1)) of L-[1-(14)C]phenylalanine. Using the breakpoint of a one-slope broken-line model, the lysine requirement was determined to be 4.88 +/- 0.96 g/kg of diet or 366 +/- 72 mg x hen(-1) x d(-1) with an upper 95% CI of 6.40 g/kg of diet or 480 mg x hen(-1) x d(-1). IAAO allows determination of individual bird amino acid requirements for specific ages and types of birds over short periods of time and enables more accurate broiler breeder pullet diet formulation.

  2. L-Methionine Production.

    PubMed

    Shim, Jihyun; Shin, Yonguk; Lee, Imsang; Kim, So Young

    2016-11-22

    L-Methionine has been used in various industrial applications such as the production of feed and food additives and has been used as a raw material for medical supplies and drugs. It functions not only as an essential amino acid but also as a physiological effector, for example, by inhibiting fat accumulation and enhancing immune response. Producing methionine from fermentation is beneficial in that microorganisms can produce L-methionine selectively using eco-sustainable processes. Nevertheless, the fermentative method has not been used on an industrial scale because it is not competitive economically compared with chemical synthesis methods. Presented are efforts to develop suitable strains, engineered enzymes, and alternative process of producing L-methionine that overcomes problems of conventional fermentation methods. One of the alternative processes is a two-step process in which the L-methionine precursor is produced by fermentation and then converted to L-methionine by enzymes. Directed efforts toward strain development and enhanced enzyme engineering will advance industrial production of L-methionine based on fermentation.

  3. Effect of dietary lysine on hepatic lysine catabolism in broilers

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Lysine is frequently a first- or second-limiting amino acid in poultry diets. Improving the efficiency of lysine use for protein synthesis would effectively lower the lysine requirement and decrease feed costs. Understanding how lysine is degraded and how the degradation is regulated would identif...

  4. Mutation of aspartic acid-351, lysine-352, and lysine-515 alters the Ca2+ transport activity of the Ca2+-ATPase expressed in COS-1 cells.

    PubMed Central

    Maruyama, K; MacLennan, D H

    1988-01-01

    Full-length cDNAs encoding neonatal and adult isoforms of the Ca2+-ATPase of rabbit fast-twitch skeletal muscle sarcoplasmic reticulum were expressed transiently in COS-1 cells. The microsomal fraction isolated from transfected COS-1 cells contained immunoreactive Ca2+-ATPase and catalyzed Ca2+ transport at rates at least 15-fold above controls. No differences were observed in either the rates or Ca2+ dependency of Ca2+ transport catalyzed by the two isoforms. Aspartic acid-351, the site of formation of the catalytic acyl phosphate in the enzyme, was mutated to asparagine, glutamic acid, serine, threonine, histidine, or alanine. In every case, Ca2+ transport activity and Ca2+-dependent phosphorylation were eliminated. Ca2+ transport was also eliminated by mutation of lysine-352 to arginine, glutamine, or glutamic acid or by mutation of Asp351-Lys352 to Lys351-Asp352. Mutation of lysine-515, the site of fluorescein isothiocyanate modification in the enzyme, resulted in diminished Ca2+ transport activity as follows: arginine, 60%; glutamine, 25%; glutamic acid, 5%. These results demonstrate the absolute requirement of acylphosphate formation for the Ca2+ transport function and define a residue important for ATP binding. They also demonstrate the feasibility of a thorough analysis of active sites in the Ca2+-ATPase by expression and site-specific mutagenesis. Images PMID:2966962

  5. Synthesis of peptides from amino acids and ATP with lysine-rich proteinoid

    NASA Technical Reports Server (NTRS)

    Nakashima, T.; Fox, S. W.

    1980-01-01

    The paper examines the synthesis of peptides from aminoacids and ATP with a lysine-rich protenoid. The latter in aqueous solution catalyzes the formation of peptides from free amino acids and ATP; this catalytic activity is not found in acidic protenoids, even though the latter contain a basic aminoacid. The pH optimum for the synthesis is about 11, but it is appreciable below 8 and above 13. Temperature data indicate an optimum at 20 C or above, with little increase in rate up to 60 C. Pyrophosphate can be used instead of ATP, but the yields are lower. The ATP-aided syntheses of peptides in aqueous solution occur with several types of proteinous aminoacids.

  6. L-lysine-L-tartaric acid: New molecular complex with nonlinear optical properties. Structure, vibrational spectra and phase transitions

    SciTech Connect

    Debrus, S.; Marchewka, M.K. . E-mail: mkm@int.pan.wroc.pl; Baran, J.; Drozd, M.; Czopnik, R.; Pietraszko, A.; Ratajczak, H.

    2005-09-15

    The first X-ray diffraction and vibrational spectroscopic analysis of a novel complex between L-lysine and L-tartaric acid is reported. The structure was solved in two temperatures (320 and 260 K) showing incommensurate phase between them. Room-temperature powder infrared and Raman measurements for the L-lysine-L-tartaric acid molecular complex (1:1) were carried out. DSC measurements on powder samples indicate two phase transitions points at about 295, 300 and 293, 300 K, for heating and cooling, respectively, with noticeable temperature interval between them. Second harmonic generation efficiency d {sub eff}=0.35 d {sub eff} (KDP)

  7. The effect of glutamic acid side chain on acidity constant of lysine in beta-sheet: A density functional theory study

    NASA Astrophysics Data System (ADS)

    Sargolzaei, M.; Afshar, M.; Sadeghi, M. S.; Kavee, M.

    2014-07-01

    In this work, the possibility of proton transfer between side chain of lysine and glutamic acid in peptide of Glu--Ala-Lys+ was demonstrated using density functional theory (DFT). We have shown that the proton transfer takes place between side chain of glutamic and lysine residues through the hydrogen bond formation. The structures of transition state for proton transfer reaction were detected in gas and solution phases. Our kinetic studies show that the proton transfer reaction rate in gas phase is higher than solution phase. The ionization constant (p K a) value of lysine residue in peptide was estimated 1.039 which is lower than intrinsic p K a of lysine amino acid.

  8. Effect on days of lactation and methionine hydroxy analog on incorporation of plasma fatty acids into plasma triglycerides

    SciTech Connect

    Pullen, D.L.; Emergy, R.S. ); Palmquist, D.L. )

    1989-01-01

    Methionine hydroxy analog has been proposed to stimulate hepatic lipoprotein synthesis and incorporation of plasma fatty acids into plasma triglyceride. Seven cows were fed diets containing 0 to 30 g analog/d starting 14 d prepartum. At approximately 30 and 60 d postpartum, cows were continuously infused intravenously with 1-({sup 14}C)palmitic acid for 160 min to achieve steady-state labeling of plasma fatty acid and triglyceride. Turnover of fatty acid and transfer quotients for triglyceride and CO{sub 2} were 3.3 an 2.7 mmol min{sup {minus}1}; 13.0 and 10.0%; and 8.0 and 5.0%, for control and analog, respectively. Proportion of fatty acid turnover incorporated into triglyceride and CO{sub 2} were 14.0 and 15.0%; and 21.0 and 18.0, respectively, for control and analog. Analog increased {sup 14}C recovered in milk fat (52 vs. 36%). Plasma concentration of fatty acids, percent oxidized to CO{sub 2}, and percent of CO{sub 2} from fatty acids decreased with increasing lactation days. Milk fat percent and yield fatty acid turnover, and oxidation were positively correlated with concentration of plasma fatty acids, whereas fatty acid incorporated into plasma triglyceride was negatively correlated with fatty acid concentration. The data suggest that hepatic triglyceride secretion is not increased in early lactation; further, no effects of analog on lipid metabolism were detected.

  9. Methionine requirements in healthy adolescents

    Technology Transfer Automated Retrieval System (TEKTRAN)

    To investigate methionine requirements in healthy adolescents, we conducted an experiment in 28 children age 15.6+2.1 years, wt. 57.8+9.7 kg, using the intravenous indicator amino acid oxidation and balance (IV-IAAOB) technique. Children were randomly assigned to 7 different levels of methionine int...

  10. Alpha-lipoic acid affects the oxidative stress in various brain structures in mice with methionine and choline deficiency

    PubMed Central

    Veskovic, Milena; Mladenovic, Dusan; Jorgacevic, Bojan; Stevanovic, Ivana; de Luka, Silvio

    2015-01-01

    Deficiency in methionine or choline can induce oxidative stress in various organs such as liver, kidney, heart, and brain. This study was to examine the effects of alpha-lipoic acid (LA) on oxidative stress induced by methionine and choline deficiency (MCD) in several brain structures. Male mice C57BL/6 (n = 28) were divided into four groups: (1) control – continuously fed with standard chow; (2) LA – fed with standard chow and receiving LA; (3) MCD2 – fed with MCD diet for two weeks, and (4) MCD2+LA – fed with MCD diet for two weeks and receiving LA (100 mg/kg/day intraperitonealy [i.p.]). Brain tissue (cortex, hypothalamus, striatum and hippocampus) was taken for determination of oxidative stress parameters. MCD diet induced a significant increase in malondialdehyde and NOx concentration in all brain regions, while LA restored their content to normal values. Similar to this, in MCD2 group, activity of total SOD, MnSOD, and Cu/ZnSOD was reduced by MCD diet, while LA treatment improved their activities in all brain structures. Besides, in MCD2 group a decrease in catalase activity in cortex and GSH content in hypothalamus was evident, while LA treatment induced an increase in catalase activity in cortex and striatum and GSH content in hypothalamus. LA treatment can significantly reduce lipid peroxidation and nitrosative stress, caused by MCD diet, in all brain regions by restoring antioxidant enzymes activities, predominantly total SOD, MnSOD, and Cu/ZnSOD, and to a lesser extent by modulating catalase activity and GSH content. LA supplementation may be used in order to prevent brain oxidative injury induced by methionine and choline deficiency. PMID:25193852

  11. Effects of dietary methionine and lysine sources on nutrient digestion, nitrogen utilization, and duodenal amino acid flow in growing goats.

    PubMed

    Sun, Z H; Tan, Z L; Liu, S M; Tayo, G O; Lin, B; Teng, B; Tang, S X; Wang, W J; Liao, Y P; Pan, Y F; Wang, J R; Zhao, X G; Hu, Y

    2007-12-01

    This study investigated the effects of supplementation of various sources of Met and Lys on nutrient digestion, N utilization, and duodenal AA flows in growing goats. Four 4-mo-old Liuyang Black wether goats were used in a 4 x 4 Latin square experiment and were assigned to 4 dietary treatments: (1) control, (2) control + lipid-coated Met-Zn chelate and Lys-Mn chelate (PML), (3) control + Met-Zn chelate and Lys-Mn chelate (CML), and (4) control + dl-Met, l-Lys-HCl, ZnSO(4).7H(2)O, and MnSO(4).H(2)O (FML). Compared with control, PML reduced (P < 0.05) ruminal NH(3) concentration, urinary N excretion, and plasma urea N concentration and increased (P < 0.05) the activity of ruminal endo-1,4-beta-d-glucanase and beta-glucosidase, the duodenal flow of N, N retention (g/d as well as % of absorbed N), the duodenal flows of Met, Lys, His, Val, and total essential AA, and plasma concentrations of Lys, Val, Phe, and total essential AA. Supplementing Zn-Met and Mn-Lys chelates had similar (P > 0.05) but lesser effects on these measures compared with PML, and the effects on most of the measures were not statistically significant (P > 0.05) when compared with control. Supplementing free-form Met and Lys had no effects compared with control (P > 0.05). The results indicate that lipid coating and chelating of AA provide a protection, and to a lesser extent by only chelating, of the AA from microbial degradation in the rumen and possibly has effects on rumen fermentation, which increases MP supply. This technology could improve productive performance and be of potential benefit to ruminant production if cost-effective products are developed.

  12. X-ray studies of crystalline complexes involving amino acids and peptides. XLIII. Adipic acid complexes of L- and DL-lysine.

    PubMed

    Sharma, Alok; Thamotharan, S; Roy, Siddhartha; Vijayan, M

    2006-03-01

    The asymmetric unit of the DL-lysine complex of adipic acid [bis(DL-lysinium) adipate], 2C6H15N2O2+.C6H8O(4)2-, contains a zwitterionic singly charged lysinium cation and half a doubly charged adipate anion (the complete anion has inversion symmetry). That of the L-lysine complex (lysinium hydrogen adipate), C6H15N2O2+.C6H9O4-, consists of a lysinium cation and a singly charged hydrogen adipate anion. In both structures, the lysinium cations organize into layers interconnected by adipate or hydrogen adipate anions. However, the arrangement of the molecular ions in the layer is profoundly different in the DL- and L-lysine complexes. The hydrogen adipate anions in the L-lysine complex form linear arrays in which adjacent ions are interconnected by a symmetric O...H...O hydrogen bond.

  13. Radical S-adenosyl methionine epimerases: regioselective introduction of diverse D-amino acid patterns into peptide natural products.

    PubMed

    Morinaka, Brandon I; Vagstad, Anna L; Helf, Maximilian J; Gugger, Muriel; Kegler, Carsten; Freeman, Michael F; Bode, Helge B; Piel, Jörn

    2014-08-04

    PoyD is a radical S-adenosyl methionine epimerase that introduces multiple D-configured amino acids at alternating positions into the highly complex marine peptides polytheonamide A and B. This novel post-translational modification contributes to the ability of the polytheonamides to form unimolecular minimalistic ion channels and its cytotoxic activity at picomolar levels. Using a genome mining approach we have identified additional PoyD homologues in various bacteria. Three enzymes were expressed in E. coli with their cognate as well as engineered peptide precursors and shown to introduce diverse D-amino acid patterns into all-L peptides. The data reveal a family of architecturally and functionally distinct enzymes that exhibit high regioselectivity, substrate promiscuity, and irreversible action and thus provide attractive opportunities for peptide engineering.

  14. Effect of amino acids lysine and arginine on fracture healing in rabbits: A radiological and histomorphological analysis

    PubMed Central

    Sinha, Shivam; Goel, Satish Chandra

    2009-01-01

    Background: Amino acids like arginine and lysine have been suggested to hasten the process of fracture healing by improving the local blood supply, supplementing growth factors, and improving collagen synthesis. We studied the role of lysine and arginine in the fracture repair process with regard to the rate of healing, probable mechanisms involved in the process, and mutual synergism between these agents. Materials and Methods: In an experimental study, 40 rabbits were subjected to ulnar osteotomy. They were distributed in control (14) and test groups (26). Twenty-six animals in the test group were fed with a diet rich in lysine and arginine. Both the groups were followed radiologically and histologically till union. Results: There was better healing of osteotomy in terms of better vascularization, callus formation, and mineralization in the test group. The time of healing in the test group was reduced by a period of 2 weeks. Conclusion: We conclude that amino acids like arginine and lysine may hasten fracture healing. PMID:19838381

  15. Nucleotide sequence of a lysine transfer ribonucleic Acid from bakers' yeast.

    PubMed

    Madison, J T; Boguslawski, S J; Teetor, G H

    1972-05-12

    The nucleotide sequence of one of the two major lysine transfer RNA's from bakers' yeast has been determined. Its structure is compared to that of a lysine tRNA from a haploid yeast. A total of 21 nucleotides differ in the two molecules. Only the T-psi-C-G (thymidine-pseudouridine-cytidine-guanosine) loop and its supporting stem are identical.

  16. The Monosodium Glutamate Story: The Commercial Production of MSG and Other Amino Acids

    NASA Astrophysics Data System (ADS)

    Ault, Addison

    2004-03-01

    Examples of the industrial synthesis of pure amino acids are presented. The emphasis is on the synthesis of ( S )-glutamic acid and, to a lesser extent, ( S )-lysine and ( R,S )-methionine. These amino acids account for about 90% of the total world production of amino acids, ( S )-glutamic acid being used as a flavor-enhancing additive (MSG) for the human diet, and ( S )-lysine and ( R,S )-methionine as supplements for the feeding of domestic animals. Examples include chemical, enzymatic, and fermentation synthesis, and two clever continuous processes for the resolution of enantiomers. See Featured Molecules .

  17. Corrosion inhibition and adsorption behavior of methionine on mild steel in sulfuric acid and synergistic effect of iodide ion.

    PubMed

    Oguzie, E E; Li, Y; Wang, F H

    2007-06-01

    The corrosion inhibition of mild steel in sulfuric acid by methionine (MTI) was investigated using electrochemical techniques. The effect of KI additives on corrosion inhibition efficiency was also studied. The results reveal that MTI inhibited the corrosion reaction by adsorption onto the metal/solution interface. Inhibition efficiency increased with MTI concentration and synergistically increased in the presence of KI, with an optimum [KI]/[MTI] ratio of 5/5, due to stabilization of adsorbed MTI cations as revealed by AFM surface morphological images. Potentiodynamic polarization data suggest that the compound functioned via a mixed-inhibition mechanism. This observation was further corroborated by the fit of the experimental adsorption data to the Temkin and Langmuir isotherms. The inhibition mechanism has been discussed vis-à-vis the presence of both nitrogen and sulfur atoms in the MTI molecule.

  18. Effect of Dietary Combination of Methionine and Fish Oil on Cellular Immunity and Plasma Fatty Acids in Infectious Bursal Disease Challenged Chickens

    PubMed Central

    Kasim, Azhar; Yong Meng, Goh; Teck Chwen, Loh; Kamalidehghan, Behnam; Soleimani Farjam, Abdoreza

    2013-01-01

    This study was carried out to investigate the modulatory effects of dietary methionine and fish oil on immune response, plasma fatty acid profile, and blood parameters of infectious bursal disease (IBD) challenged broiler chickens. A total of 300 one-day-old male broiler chicks were assigned to one of six dietary treatment groups in a 3 × 2 factorial arrangement. There were three levels of fish oil (0, 2.5 and 5.5%), and two levels of methionine (NRC recommendation and twice NRC recommendation). The results showed that the birds fed with 5.5% fish oil had higher total protein, white blood cell count, and IL-2 concentration than those of other groups at 7 days after IBD challenge. Inclusion of fish oil in diet had no effect on IFN-γ concentration. However, supplementation of methionine twice the recommendation enhanced the serum IFN-γ and globulin concentration. Neither of fish oil nor methionine supplementation affected the liver enzymes concentration. It can be suggested that a balance of moderate level of fish oil (2.5%) and methionine level (twice NRC recommendation) might enhance immune response in IBD challenged broiler chickens. PMID:24198724

  19. Effect of dietary combination of methionine and fish oil on cellular immunity and plasma fatty acids in infectious bursal disease challenged chickens.

    PubMed

    Maroufyan, Elham; Kasim, Azhar; Yong Meng, Goh; Ebrahimi, Mahdi; Teck Chwen, Loh; Mehrbod, Parvaneh; Kamalidehghan, Behnam; Soleimani Farjam, Abdoreza

    2013-01-01

    This study was carried out to investigate the modulatory effects of dietary methionine and fish oil on immune response, plasma fatty acid profile, and blood parameters of infectious bursal disease (IBD) challenged broiler chickens. A total of 300 one-day-old male broiler chicks were assigned to one of six dietary treatment groups in a 3 × 2 factorial arrangement. There were three levels of fish oil (0, 2.5 and 5.5%), and two levels of methionine (NRC recommendation and twice NRC recommendation). The results showed that the birds fed with 5.5% fish oil had higher total protein, white blood cell count, and IL-2 concentration than those of other groups at 7 days after IBD challenge. Inclusion of fish oil in diet had no effect on IFN- γ concentration. However, supplementation of methionine twice the recommendation enhanced the serum IFN- γ and globulin concentration. Neither of fish oil nor methionine supplementation affected the liver enzymes concentration. It can be suggested that a balance of moderate level of fish oil (2.5%) and methionine level (twice NRC recommendation) might enhance immune response in IBD challenged broiler chickens.

  20. Metabolism of lysine in alpha-aminoadipic semialdehyde dehydrogenase-deficient fibroblasts: evidence for an alternative pathway of pipecolic acid formation.

    PubMed

    Struys, Eduard A; Jakobs, Cornelis

    2010-01-04

    The mammalian degradation of lysine is believed to proceed via two distinct routes, the saccharopine and the pipecolic acid routes, that ultimately converge at the level of alpha-aminoadipic semialdehyde (alpha-AASA). alpha-AASA dehydrogenase-deficient fibroblasts were grown in cell culture medium supplemented with either L-[alpha-(15)N]lysine or L-[epsilon-(15)N]lysine to explore the exact route of lysine degradation. L-[alpha-(15)N]lysine was catabolised into [(15)N]saccharopine, [(15)N]alpha-AASA, [(15)N]Delta(1)-piperideine-6-carboxylate, and surprisingly in [(15)N]pipecolic acid, whereas L-[epsilon-(15)N]lysine resulted only in the formation of [(15)N]saccharopine. These results imply that lysine is exclusively degraded in fibroblasts via the saccharopine branch, and pipecolic acid originates from an alternative precursor. We hypothesize that pipecolic acid derives from Delta(1)-piperideine-6-carboxylate by the action of Delta(1)-pyrroline-5-carboxylic acid reductase, an enzyme involved in proline metabolism.

  1. Identification of Structural and Catalytic Classes of Highly Conserved Amino Acid Residues in Lysine 2,3-Aminomutase †

    PubMed Central

    Chen, Dawei; Frey, Perry A.; Lepore, Bryan W.; Ringe, Dagmar; Ruzicka, Frank J.

    2008-01-01

    Lysine 2,3-aminomutase (LAM) from Clostridium subterminale SB4 catalyzes the interconversion of (S)-lysine and (S)-β-lysine by a radical mechanism involving coenzymatic actions of S-adenosylmethionine (SAM), a [4Fe-4S] cluster, and pyridoxal-5′-phosphate (PLP). The enzyme contains a number of conserved acidic residues and a cysteine and arginine-rich motif, that binds iron and sulfide in the [4Fe–4S] cluster. The results of activity and iron, sulfide, and PLP analysis of variants resulting from site-specific mutations of the conserved acidic residues and the arginine residues in the iron-sulfide binding motif indicate two classes of conserved residues of each type. Mutation of the conserved residues Arg134, Asp293, and Asp330 abolish all enzymatic activity. Based on the x-ray crystal structure, these residues bind the ε-aminium and α-carboxylate groups of (S)-lysine. However, among these residues only Asp293 appears to be important for stabilizing the [4Fe–4S] cluster. Members of a second group of conserved residues appear to stabilize the structure of LAM. Mutations of arginine residues 130, 135, and 136 and acidic residues Glu86, Asp165, Glu236, and Asp172 dramatically decrease iron and sulfide contents in the purified variants. Mutation of Asp96 significantly decreases iron and sulfide content. Variants in Arg130 or Asp172 display no detectable activity, whereas variants in the other positions display low to very low activities. Structural roles are assigned to this latter class of conserved amino acids. In particular, a network of hydrogen bonded interactions of Arg130, Glu86, Arg135 and the main chain carbonyl groups of Cys132 and Leu55 appears to stabilize the [4Fe–4S] cluster. PMID:17042481

  2. Antiapoptotic role of S-adenosyl-l-methionine against hydrochloric acid induced cell death in Saccharomyces cerevisiae.

    PubMed

    Malakar, Dipankar; Dey, Anindya; Basu, Arghya; Ghosh, Anil K

    2008-01-01

    Exposure of stationary phase cells of Saccharomyces cerevisiae to 10 mM HCl (pH approximately 2) resulted in cell death as a function of time (up to 6 h) with most (about 40%-65%) of the cells showing apoptotic features including chromatin condensation along the nuclear envelope, exposure of phosphatidylserine on the outer leaflet of cytoplasmic membrane, and DNA fragmentation. During the first 2 h of acid exposure there was an increase in reactive oxygen species (ROS) level inside cells, with subsequent elevation in the level of lipid peroxidation and decrease in reducing equivalents culminating in loss of mitochondrial membrane potential (DeltaPsi(m)). An initial (1 h) event of mitochondrial hyper-polarization with subsequent elevation of ROS level of the acid treated cells was also observed. S-adenosyl-l-methionine (AdoMet; 1 mM) treatment increased the cell survival of the acid stressed cells. It partially scavenged the increased intracellular ROS level by supplementing glutathione through the transsulfuration pathway. It also inhibited acid mediated lipid peroxidation, partially recovered acid evoked loss of DeltaPsi(m) and protected the cells from apoptotic cell death. S-adenosyl di-aldehyde, an indirect inhibitor of the AdoMet metabolic pathway, increased mortality of the acid treated cells. Incubation of acid stressed cells with the antioxidant, N-acetyl-cysteine (1 mM), decreased the cellular mortality, but the same concentration of AdoMet offered more protection by scavenging the free radicals. The ability of AdoMet to scavenge ROS mediated apoptosis may be an important function of this molecule in responding to cellular stress. The study could open a new avenue for detailed investigation on the curative potential of AdoMet against gastric ulcer.

  3. Synthesis of stereoarray isotope labeled (SAIL) lysine via the "head-to-tail" conversion of SAIL glutamic acid.

    PubMed

    Terauchi, Tsutomu; Kamikawai, Tomoe; Vinogradov, Maxim G; Starodubtseva, Eugenia V; Takeda, Mitsuhiro; Kainosho, Masatsune

    2011-01-07

    A stereoarray isotope labeled (SAIL) lysine, (2S,3R,4R,5S,6R)-[3,4,5,6-(2)H(4);1,2,3,4,5,6-(13)C(6);2,6-(15)N(2)]lysine, was synthesized by the "head-to-tail" conversion of SAIL-Glu, (2S,3S,4R)-[3,4-(2)H(2);1,2,3,4,5-(13)C(5);2-(15)N]glutamic acid, with high stereospecificities for all five chiral centers. With the SAIL-Lys in hand, the unambiguous simultaneous stereospecific assignments were able to be established for each of the prochiral protons within the four methylene groups of the Lys side chains in proteins.

  4. Use of the guanidination reaction for determining reactive lysine, bioavailable lysine and gut endogenous lysine.

    PubMed

    Rutherfurd, Shane M

    2015-09-01

    Determining the bioavailability of lysine in foods and feedstuffs is important since lysine is often the first limiting indispensable amino acid in diets for intensively farmed livestock (pigs and poultry) and also in many cereal-based diets consumed by humans. When foods or feedstuffs are heat processed, lysine can undergo Maillard reactions to produce nutritionally unavailable products. The guanidination reaction, the reaction of O-methylisourea with the side chain amino group of lysine that produces homoarginine, has been used to determine the unmodified lysine (reactive lysine) in processed foods and feedstuffs and also true ileal digestible reactive lysine (bioavailable lysine). The advantages of the guanidination method in comparison with other reactive lysine methods such as the fluorodinitrobenzene, trinitrobenzenesulphonic acid and dye-binding methods are that it is very specific for reactive lysine and also that the method is relatively straightforward to conduct. The specificity of the guanidination reaction for the lysine side chain amino group is particularly important, since ileal digesta will contain N-terminal groups in the form of free amino acids and peptides. The main disadvantage is that complete conversion of lysine to homoarginine is required, yet it is not straightforward to test for complete guanidination in processed foods and feedstuffs. Another disadvantage is that the guanidination reaction conditions may vary for different food types and sometimes within the same food type. Consequently, food-specific guanidination reaction conditions may be required and more work is needed to optimise the reaction conditions across different foods and feedstuffs.

  5. Influence of copper supplementation on the relationship between dietary methionine and free plasma methionine.

    PubMed

    Ekperigin, H E; Vohra, P

    1981-09-01

    One-day-old broiler chicks were adapted to a basal, isolated soyprotein-cornstarch diet containing 20% protein, 0.59% methionine and 3,300 kcal metabolizable energy (ME)/kg. They were then fed experimental diets consisting of three levels (0, 500, 1,000 ppm) of copper added to each of the basal diet plus four levels (0, 0.4, 0.8, or 1.5%) of L-methionine for 1 or 4 weeks. Growth was retarded after 1 week by all levels of supplementary methionine, and by 500 or 1,000 ppm excess copper. The plasma concentrations of free methionine, serine, alpha-aminobutyric acid and cystathionine were increased by excess dietary methionine. Excess copper prevented the increase in plasma methionine. After 4 weeks, the plasma methionine concentration and rate of growth of chicks fed 0.4% excess L-methionine did not differ significantly from basal values. The growth retardation caused by 500 ppm excess copper was alleviated by 0.4% supplemental methionine, and the elevations in plasma methionine and liver and spleen iron concentrations observed in chicks fed 1.5% excess methionine were reduced by 1,000 ppm excess copper. The patterns of the relationship between dietary methionine and liver or spleen iron, in the presence or absence of supplementary copper, were similar to those between dietary and free plasma methionine.

  6. Lysine biosynthesis and nitrogen metabolism in quinoa (Chenopodium quinoa): study of enzymes and nitrogen-containing compounds.

    PubMed

    Varisi, Vanderlei A; Camargos, Liliane S; Aguiar, Leandro F; Christofoleti, Renata M; Medici, Leonardo O; Azevedo, Ricardo A

    2008-01-01

    Aspartate kinase (AK, EC 2.7.2.4), homoserine dehydrogenase (HSDH, EC 1.1.1.3) and dihydrodipicolinate synthase (DHDPS, EC 4.2.1.52) were isolated and partially purified from immature Chenopodium quinoa Willd seeds. Enzyme activities were studied in the presence of the aspartate-derived amino acids lysine, threonine and methionine and also the lysine analogue S-2-aminoethyl-l-cysteine (AEC), at 1 mM and 5 mM. The results confirmed the existence of, at least, two AK isoenzymes, one inhibited by lysine and the other inhibited by threonine, the latter being predominant in quinoa seeds. HSDH activity was also shown to be partially inhibited by threonine, whereas some of the activity was resistant to the inhibitory effect, indicating the presence of two isoenzymes, one resistant and another sensitive to threonine inhibition. Only one DHDPS isoenzyme highly sensitive to lysine inhibition was detected. The results suggest that the high concentration of lysine observed in quinoa seeds is possibly due to a combined effect of increased lysine synthesis and accumulation in the soluble form and/or as protein lysine. Nitrogen assimilation was also investigated and based on nitrate content, nitrate reductase activity, amino acid distribution and ureide content, the leaves were identified as the predominant site of nitrate reduction in this plant species. The amino acid profile analysis in leaves and roots also indicated an important role of soluble glutamine as a nitrogen transporting compound.

  7. Methionine requirements for the preimplantation bovine embryo.

    PubMed

    BONILLA, Luciano; LUCHINI, Daniel; DEVILLARD, Estelle; HANSEN, Peter J

    2010-10-01

    The early embryo's nutritional environment plays an important role in establishing its developmental potential. However, little is known about the specific nutrient requirements of the embryo. The objective of the present study was to determine requirements of the in vitro produced bovine embryo for the essential amino acid methionine. In addition to serving as a precursor for polypeptides, methionine plays roles in regulation of translation, DNA methylation, and antioxidant balance. In the first experiment, embryos were cultured in potassium simplex optimized medium - bovine embryo modification 2 containing 0, 35, 50, 100, 200 or 400 µmol/l L-methionine for 8 days. There was no effect of methionine concentration on cleavage rate. The percent of oocytes that developed to blastocyst was lower for embryos without methionine at Day 7 and 8 than other groups but was similar for embryos cultured with 35-400 µmol/l. Neither total cell number, allocation of cells to trophectoderm or inner cell mass, or frequency of apoptosis was affected by methionine concentration. In the second experiment, embryos were cultured with 0, 7, 14, 21, 28 or 35 µmol/l methionine. There was no effect of methionine concentration on cleavage rate. The percent of oocytes that developed to blastocyst was lower for embryos without methionine at Day 7 and 8 but was not different between embryos cultured with 7-35 µmol/l methionine. However, the proportion of blastocysts that were expanded, hatching or hatched on Day 7 was reduced at lower concentrations of methionine (7 and 14). DNA methylation of blastocyst nuclei was unaffected by methionine concentration but intracellular glutathione content was higher for embryos cultured without methionine. In conclusion, the methionine requirement for preimplantation development is between 14 and 21 µmol/l. These concentrations are lower or similar to those found in the reproductive tract and suggest that methionine deficiency is not a common cause of

  8. The effect of dietary methionine levels on endogenous nitrogen and endogenous amino acids flows in growing goats.

    PubMed

    Zhou, C S; Tan, Z L; Tang, S X; Sun, Z H; Han, X F; Wang, M; Tayo, G O

    2010-10-01

    The effect of dietary methionine (Met) levels on endogenous N and amino acids (AA) flows at different part of the digestive tract of growing goats was determined using a (15)N isotope dilution technique. Three goats (25 ± 2.5 kg) were fitted with the ruminal, duodenal and ileal cannulae and allocated to three dietary treatments in a 3 × 3 Latin square design. The dietary treatments consisted of a total mixed ration containing three levels of Met (0.15%, 0.25% and 0.35%) respectively. It was found that at 0.15% Met level, the lowest flow in endogenous N and total AA at the duodenum and ileum occurred. The endogenous N secretion contributed to 26% and 23% of the duodenal and ileal total N flows, respectively, and the proportions were not affected by the dietary Met levels. The duodenal and ileal flows of endogenous total AA were 11.1, 11.8, 11.3 g/d and 2.9, 3.9, 4.1 g/d respectively. The average real digestibility of N was 65%, 87% and 95% in the forestomach, intestine and whole digestive tract respectively.

  9. L-methionine-dl-sulfoxide metabolism and toxicity in freshly isolated mouse hepatocytes: gender differences and inhibition with aminooxyacetic acid.

    PubMed

    Dever, Joseph T; Elfarra, Adnan A

    2008-11-01

    L-methionine-dl-sulfoxide (MetO) is an L-methionine (Met) metabolite, but its role in Met metabolism and toxicity is not clear. In this study, MetO uptake, metabolism to Met, cytotoxicity, and glutathione (GSH) and glutathione disulfide (GSSG) status were characterized in freshly isolated mouse hepatocytes incubated at 37 degrees C with 0 to 30 mM MetO for 0 to 5 h. In male hepatocytes, dose-dependent cytotoxicity concomitant with GSH depletion without GSSG formation occurred after exposure to 20 or 30 mM MetO but not after exposure to 10 mM MetO. Interestingly, female hepatocytes exposed to 30 mM MetO showed no cytotoxicity and exhibited increased intracellular GSH levels compared with control hepatocytes. Male hepatocytes had approximately 2-fold higher levels of intracellular Met-d-O or Met-l-O after MetO (30 mM) exposure for 0 to 1.5 h compared with female hepatocytes. In hepatocytes of both genders, Met-l-O was detected at nearly 5-fold higher levels than Met-d-O, and no significant increase in cellular Met levels was detected. Addition of aminooxyacetic acid (AOAA), an inhibitor of transamination reactions, to MetO-exposed male hepatocytes resulted in higher cellular Met-d-O and Met-l-O levels and decreased the cytotoxicity of MetO. Interestingly, exposure of control male hepatocytes to AOAA selectively increased cellular Met-d-O levels to levels similar to those observed after exposure to MetO (30 mM). Analysis of MetO transamination activity by glutamine transaminase K in mouse liver cytosol revealed similar rates of MetO transamination in cytosol of both genders. Taken together, these results provide evidence for stereoselective oxidation of Met to Met-d-O under physiological conditions and suggest a major role for MetO transamination in MetO metabolism and toxicity.

  10. Lysine-functionalized nanodiamonds: synthesis, physiochemical characterization, and nucleic acid binding studies

    PubMed Central

    Kaur, Randeep; Chitanda, Jackson M; Michel, Deborah; Maley, Jason; Borondics, Ferenc; Yang, Peng; Verrall, Ronald E; Badea, Ildiko

    2012-01-01

    Purpose: Detonation nanodiamonds (NDs) are carbon-based nanomaterials that, because of their size (4–5 nm), stable inert core, alterable surface chemistry, fluorescence, and biocompatibility, are emerging as bioimaging agents and promising tools for the delivery of biochemical molecules into cellular systems. However, diamond particles possess a strong propensity to aggregate in liquid formulation media, restricting their applicability in biomedical sciences. Here, the authors describe the covalent functionalization of NDs with lysine in an attempt to develop nanoparticles able to act as suitable nonviral vectors for transferring genetic materials across cellular membranes. Methods: NDs were oxidized and functionalized by binding lysine moieties attached to a three-carbon-length linker (1,3-diaminopropane) to their surfaces through amide bonds. Raman and Fourier transform infrared spectroscopy, zeta potential measurement, dynamic light scattering, atomic force microscopic imaging, and thermogravimetric analysis were used to characterize the lysine-functionalized NDs. Finally, the ability of the functionalized diamonds to bind plasmid DNA and small interfering RNA was investigated by gel electrophoresis assay and through size and zeta potential measurements. Results: NDs were successfully functionalized with the lysine linker, producing surface loading of 1.7 mmol g−1 of ND. These modified NDs formed highly stable aqueous dispersions with a zeta potential of 49 mV and particle size of approximately 20 nm. The functionalized NDs were found to be able to bind plasmid DNA and small interfering RNA by forming nanosized “diamoplexes”. Conclusion: The lysine-substituted ND particles generated in this study exhibit stable aqueous formulations and show potential for use as carriers for genetic materials. PMID:22904623

  11. The physiological role of reversible methionine oxidation.

    PubMed

    Drazic, Adrian; Winter, Jeannette

    2014-08-01

    Sulfur-containing amino acids such as cysteine and methionine are particularly vulnerable to oxidation. Oxidation of cysteine and methionine in their free amino acid form renders them unavailable for metabolic processes while their oxidation in the protein-bound state is a common post-translational modification in all organisms and usually alters the function of the protein. In the majority of cases, oxidation causes inactivation of proteins. Yet, an increasing number of examples have been described where reversible cysteine oxidation is part of a sophisticated mechanism to control protein function based on the redox state of the protein. While for methionine the dogma is still that its oxidation inhibits protein function, reversible methionine oxidation is now being recognized as a powerful means of triggering protein activity. This mode of regulation involves oxidation of methionine to methionine sulfoxide leading to activated protein function, and inactivation is accomplished by reduction of methionine sulfoxide back to methionine catalyzed by methionine sulfoxide reductases. Given the similarity to thiol-based redox-regulation of protein function, methionine oxidation is now established as a novel mode of redox-regulation of protein function. This article is part of a Special Issue entitled: Thiol-Based Redox Processes.

  12. Novel sirtuin inhibitory warheads derived from the N(ε)-acetyl-lysine analog L-2-amino-7-carboxamidoheptanoic acid.

    PubMed

    He, Yanhua; Yan, Lingling; Zang, Wenwen; Zheng, Weiping

    2015-11-14

    Built upon the catalytic mechanism-based pan-SIRT1/2/3 inhibitory warhead L-2-amino-7-carboxamidoheptanoic acid (L-ACAH, a close structural analog of N(ε)-acetyl-lysine) that our laboratory discovered recently, in the current study, its carboxamide NH2-ethylated analog was found to be a ∼2.4-6.6-fold stronger SIRT1/2/3 inhibitory warhead than L-ACAH. Carboxamide NH2-dodecylated and carboxymethylated analogs of L-ACAH were also identified as potent SIRT6 and SIRT5 inhibitory warheads, respectively.

  13. Investigation of nonfouling polypeptides of poly(glutamic acid) with lysine side chains synthesized by EDC·HCl/HOBt chemistry.

    PubMed

    Yang, Qinghua; Li, Wenchen; Wang, Longgang; Wang, Guangzhi; Wang, Zhen; Liu, Lingyun; Chen, Shengfu

    2014-01-01

    Nonfouling polypeptides with homogenous alternating charges draw peoples' attentions for their potential capability in biodegradation. Homogenous glutamic acid (E) and lysine (K) polypeptides were proposed and synthesized before. In this work, a new polypeptide formed by poly(glutamic acid) with lysine side chains (poly(E)-K) was synthesized by facile EDC·HCl/HOBt chemistry and investigated. Results show that these polypeptides also have good nonspecific protein resistance determined by enzyme-linked immunosorbent assay. The lowest nonspecific adsorption of the model proteins, anti-IgG and fibrinogen (Fg), on the self-assembling monolayers (SAMs) surface of poly(E)-K was only 3.3 ± 1.8 and 4.4 ± 1.6%, respectively, when protein adsorption on tissue culture polystyrene surface was set as 100%. And, the relative nonspecific protein adsorption increases when the polypeptide molecular weight increases due to the repression of low density polymer brushes. Moreover, almost no obvious cytotoxicity and hemolytic activity in vitro were detected. This work suggests that polypeptides with various formats of homogenous balanced charges could achieve excellent nonspecific protein resistance, which might be the intrinsic reason for the coexistence of high concentration serum proteins in blood.

  14. THE BASIS OF STABILITY IN LYSINE AND ARGININE SALTS OF UNSATURATED FATTY ACIDS.

    DTIC Science & Technology

    LINOLEIC ACID , STABILIZATION), (* FATTY ACIDS , STABILITY), (*AMINO ACIDS , SALTS), (*ANTIOXIDANTS, AMINO ACIDS ), DEHYDRATED FOODS, ADDITIVES...PRESERVATION, COMPLEX COMPOUNDS, ELECTRICAL CONDUCTIVITY, INFRARED SPECTRA, NUCLEAR MAGNETIC RESONANCE, CHROMATOGRAPHIC ANALYSIS, X RAY DIFFRACTION, CRYSTAL LATTICES, MOLECULAR ISOMERISM, FATTY ACID ESTERS

  15. Metabolic pathways promoting intrahepatic fatty acid accumulation in methionine and choline deficiency: implications for the pathogenesis of steatohepatitis.

    PubMed

    Macfarlane, David P; Zou, Xiantong; Andrew, Ruth; Morton, Nicholas M; Livingstone, Dawn E W; Aucott, Rebecca L; Nyirenda, Moffat J; Iredale, John P; Walker, Brian R

    2011-02-01

    The pathological mechanisms that distinguish simple steatosis from steatohepatitis (or NASH, with consequent risk of cirrhosis and hepatocellular cancer) remain incompletely defined. Whereas both a methionine- and choline-deficient diet (MCDD) and a choline-deficient diet (CDD) lead to hepatic triglyceride accumulation, MCDD alone is associated with hepatic insulin resistance and inflammation (steatohepatitis). We used metabolic tracer techniques, including stable isotope ([¹³C₄]palmitate) dilution and mass isotopomer distribution analysis (MIDA) of [¹³C₂]acetate, to define differences in intrahepatic fatty acid metabolism that could explain the contrasting effect of MCDD and CDD on NASH in C57Bl6 mice. Compared with control-supplemented (CS) diet, liver triglyceride pool sizes were similarly elevated in CDD and MCDD groups (24.37 ± 2.4, 45.94 ± 3.9, and 43.30 ± 3.5 μmol/liver for CS, CDD, and MCDD, respectively), but intrahepatic neutrophil infiltration and plasma alanine aminotransferase (31 ± 3, 48 ± 4, 231 ± 79 U/l, P < 0.05) were elevated only in MCDD mice. However, despite loss of peripheral fat in MCDD mice, neither the rate of appearance of palmitate (27.2 ± 3.5, 26.3 ± 2.3, and 28.3 ± 3.5 μmol·kg⁻¹·min⁻¹) nor the contribution of circulating fatty acids to the liver triglyceride pool differed between groups. Unlike CDD, MCDD had a defect in hepatic triglyceride export that was confirmed using intravenous tyloxapol (142 ± 21, 122 ± 15, and 80 ± 7 mg·kg⁻¹·h⁻¹, P < 0.05). Moreover, hepatic de novo lipogenesis was significantly elevated in the MCDD group only (1.4 ± 0.3, 2.3 ± 0.4, and 3.4 ± 0.4 μmol/day, P < 0.01). These findings suggest that important alterations in hepatic fatty acid metabolism may promote the development of steatohepatitis. Similar mechanisms may predispose to hepatocyte damage in human NASH.

  16. Modulation of Potassium Channel Function by Methionine Oxidation and Reduction

    NASA Astrophysics Data System (ADS)

    Ciorba, Matthew A.; Heinemann, Stefan H.; Weissbach, Herbert; Brot, Nathan; Hoshi, Toshinori

    1997-09-01

    Oxidation of amino acid residues in proteins can be caused by a variety of oxidizing agents normally produced by cells. The oxidation of methionine in proteins to methionine sulfoxide is implicated in aging as well as in pathological conditions, and it is a reversible reaction mediated by a ubiquitous enzyme, peptide methionine sulfoxide reductase. The reversibility of methionine oxidation suggests that it could act as a cellular regulatory mechanism although no such in vivo activity has been demonstrated. We show here that oxidation of a methionine residue in a voltage-dependent potassium channel modulates its inactivation. When this methionine residue is oxidized to methionine sulfoxide, the inactivation is disrupted, and it is reversed by coexpression with peptide methionine sulfoxide reductase. The results suggest that oxidation and reduction of methionine could play a dynamic role in the cellular signal transduction process in a variety of systems.

  17. Effect of L- or DL-methionine Supplementation on Nitrogen Retention, Serum Amino Acid Concentrations and Blood Metabolites Profile in Starter Pigs

    PubMed Central

    Tian, Q. Y.; Zeng, Z. K.; Zhang, Y. X.; Long, S. F.; Piao, X. S.

    2016-01-01

    The objective of the current study was to evaluate the effect of supplementation of either L-methionine (L-Met) or DL-methionine (DL-Met) to diets of starter pigs on nitrogen (N) balance, metabolism, and serum amino acid profile. Eighteen crossbred (Duroc×Landrace×Yorkshire) barrows weighing 15.45±0.88 kg were randomly allotted to 1 of 3 diets with 6 pigs per treatment. The diets included a basal diet (Met-deficient diet) containing 0.24% standardized ileal digestibility Met with all other essential nutrients meeting the pig’s requirements. The other two diets were produced by supplementing the basal diet with 0.12% DL-Met or L-Met. The experiment lasted for 18 days, consisting of a 13-day adaptation period to the diets followed by a 5-day experimental period. Pigs were fed ad libitum and free access to water throughout the experiment. Results showed that the supplementation of either L-Met or DL-Met improved N retention, and serum methionine concentration, and decreased N excretion compared with basal diet (p<0.01). The N retention of pigs fed diets supplemented with the same inclusion levels of DL-Met or L-Met were not different (p>0.05). In conclusion, on equimolar basis DL-Met and L-Met are equally bioavailable as Met sources for starter pigs. PMID:26954214

  18. Infrared and Raman spectroscopy and DFT calculations of DL amino acids: Valine and lysine hydrochloride

    NASA Astrophysics Data System (ADS)

    Paiva, F. M.; Batista, J. C.; Rêgo, F. S. C.; Lima, J. A.; Freire, P. T. C.; Melo, F. E. A.; Mendes Filho, J.; de Menezes, A. S.; Nogueira, C. E. S.

    2017-01-01

    Single crystals of DL-valine and DL-lysine hydrochloride were grown by slow evaporation method and the crystallographic structure were confirmed by X-ray diffraction experiment and Rietveld method. These two crystals have been studied by Raman spectroscopy in the 25-3600 cm-1 spectral range and by infrared spectroscopy through the interval 375-4000 cm-1 at room temperature. Experimental and theoretical vibrational spectra were compared and a complete analysis of the modes was done in terms of the Potential Energy Distribution (PED).

  19. Effects of adding liquid DL-methionine hydroxy analogue-free acid to drinking water on growth performance and small intestinal morphology of nursery pigs.

    PubMed

    Kaewtapee, C; Krutthai, N; Poosuwan, K; Poeikhampha, T; Koonawootrittriron, S; Bunchasak, C

    2010-06-01

    This study was conducted to evaluate the effect of adding liquid DL-methionine hydroxy analogue free acid (LMA) to drinking water on growth performance, small intestinal morphology and volatile fatty acids in the caecum of nursery pigs. Twenty-four crossbred pigs (Large White x Landrace, BW approximately 18 kg) were divided into three groups with four replications of two piglets each. The piglets received drinking water without (control), with 0.05 or 0.10% LMA. The results indicated that adding LMA at 0.10% to drinking water significantly increased their weight gain, average daily feed intake (p < 0.05) and tended to improve the feed conversion ratio. Adding LMA to drinking water significantly increased their water intake and significantly reduced the pH of drinking water (p < 0.01), thus total plate count (p < 0.01) and Escherichia coli in drinking water was reduced (p < 0.05), while the total number of bacteria in the caecum was not significantly affected. Liquid DL-methionine hydroxy analogue free acid supplementation in drinking water tended to decrease pH in the stomach, duodenum, jejunum, colon and rectum. Furthermore, adding LMA at 0.10% significantly increased villous height in the duodenum, jejunum and ileum (p < 0.05), and the villous height:crypt depth ratio in the jejunum and ileum (p < 0.01) was higher, whereas acetic acid concentration in the caecum was significantly lower than in the control group. It could be concluded that adding LMA to drinking water improved growth performance of the nursery pigs because of high water quality and high nutrient utilization caused by an improvement of small intestinal morphology (not from nutritional effect of methionine source).

  20. Effects of intramuscular injections of vitamin B12 on lactation performance of dairy cows fed dietary supplements of folic acid and rumen-protected methionine.

    PubMed

    Girard, C L; Matte, J J

    2005-02-01

    The experiment was undertaken to determine the effects of i.m. injections of vitamin B(12) on lactational performance of primiparous dairy cows fed dietary supplements of folic acid and rumen-protected methionine from 4 to 18 wk of lactation. Fourteen primiparous Holstein cows were assigned to 7 blocks of 2 cows each, according to milk production during the third week of lactation. All cows were fed a basal diet supplemented daily with rumen-protected methionine (18 g of supplement, to bring the estimated supply of methionine to 2.2% of metabolizable protein) plus folic acid (4 mg per kg of BW). Within each block, the cows received a weekly i.m. injection (2 mL) of saline or 10 mg of vitamin B(12). Milk production was recorded daily. Milk and blood were sampled every 2 wk. Supplementary vitamin B(12) increased energy-corrected milk from 25.8 to 29.0 (SE 1.6) kg/d, as well as milk yields of solids [3.52 to 3.90 (SE 0.22) kg/d], fat [0.87 to 1.01 (SE 0.06) kg/d], and lactose [1.48 to 1.64 (SE 0.11) kg/d]. Supplementation also increased concentrations and amounts of vitamin B(12) secreted in milk but had no significant effect on dry matter intake and concentrations and amounts of folates in milk. Packed cell volume, blood hemoglobin, and serum vitamin B(12) were increased by supplementary vitamin B(12), whereas serum methylmalonic acid was decreased. Serum concentrations of sulfur amino acids were unchanged by treatment. These findings support the hypothesis that, in early lactation, supply of vitamin B(12) was not optimal and limited the lactation performance of the cows.

  1. Evaluation of amino acids as turfgrass nematicides.

    PubMed

    Zhang, Yun; Luc, John E; Crow, William T

    2010-12-01

    Laboratory experiments revealed that DL-methionine, sodium methionate, potassium methionate, and methionine hydroxyl analog at rates of 224 and 448 kg amino acid/ha reduced the number of Belonolaimus longicaudatus mixed life-stages and Meloidogyne incognita J2 in soil, whereas L-threonine and lysine were not effective in reducing the number of either nematode. Futhermore, greenhouse experiments demonstrated that DL-methionine, sodium methionate, potassium methionate, and methionine hydroxyl analog were equally effective against B. longicaudatus at rates of 112, 224, and 448 kg amino acid/ha, and the highest rate (448 kg amino acid/ha) of all amino acids was more effective in reducing the number of B. longicaudatus than the lower rate. However, phytotoxicity was observed on creeping bentgrass (Agrostis palustris) treated with 448 kg amino acid/ha of methionine hydroxyl analog and DL methionine. In addition, in one of two field experiments on bermudagrass (Cynodon dactylon × C. transvaalensis) turf percentage green cover was increased and the number of B. longicaudatus was reduced by 224 kg amino acid/ha of DL-methionine and potassium methionate compared to untreated controls in one of two trials.

  2. Preparation, characterization, and biocompatibility evaluation of poly(Nɛ-acryloyl-L-lysine)/hyaluronic acid interpenetrating network hydrogels.

    PubMed

    Cui, Ning; Qian, Junmin; Xu, Weijun; Xu, Minghui; Zhao, Na; Liu, Ting; Wang, Hongjie

    2016-01-20

    In the present study, poly(Nɛ-acryloyl-L-lysine)/hyaluronic acid (pLysAAm/HA) interpenetrating network (IPN) hydrogels were successfully fabricated through the combination of hydrazone bond crosslinking and photo-crosslinking reactions. The HA hydrogel network was first synthesized from 3,3'-dithiodipropionate hydrazide-modified HA and polyethylene glycol dilevulinate by hydrazone bond crosslinking. The pLysAAm hydrogel network was prepared from Nɛ-acryloyl-L-lysine and N,N'-bis(acryloyl)-(L)-cystine by photo-crosslinking. The resultant pLysAAm/HA hydrogels had a good shape recovery property after loading and unloading for 1.5 cycles (up to 90%) and displayed a highly porous microstructure. Their compressive moduli were at least 5 times higher than that of HA hydrogels. The pLysAAm/HA hydrogels had an equilibrium swelling ratio of up to 37.9 and displayed a glutathione-responsive degradation behavior. The results from in vitro biocompatibility evaluation with pre-osteoblasts MC3T3-E1 cells revealed that the pLysAAm/HA hydrogels could support cell viability and proliferation. Hematoxylin and eosin staining indicated that the pLysAAm/HA hydrogels allowed cell and tissue infiltration, confirming their good in vivo biocompatibility. Therefore, the novel pLysAAm/HA IPN hydrogels have great potential for bone tissue engineering applications.

  3. A novel transdermal fomulation of 18β-glycyrrhetic acid with lysine for improving bioavailability and efficacy.

    PubMed

    Li, S; Qiu, Y Q; Zhang, S H; Gao, Y H

    2012-01-01

    The aim of this study was to develop a novel topical liposomal system entrapping 18β-glycyrrhetic acid (GA) for the treatment of chronic allergic dermatitis. A novel liposomal system with molar ratios of GA to lysine from 1:1 to 1:3 was prepared by high-pressure homogenization. The liposomes at the optimized molar ratio of GA to lysine of 1:2 significantly improved GA loading (1.2%) and penetration in vitro. Liposomal gels containing GA 0.3, 0.6 and 0.9% were prepared to investigate the dosage effect on transdermal delivery and anti-inflammatory activity. In vivo pharmacokinetic studies were carried out by nonocclusive application of GA liposomal gels to ICR mouse ears. GA concentrations in skin and plasma increased proportionally with dose over the dose range of 0.3-0.9%. A 2,4-dinitrofluorobenzene-induced contact dermatitis mouse model was made up to evaluate the pharmacodynamics of GA liposomal gels. The liposomal gel with GA 0.9% showed a stronger anti-inflammatory activity than triamcinolone acetonide and econazole nitrate cream, while few side effects were observed in the present model. The topical administration of gel containing novel elastic liposomes of GA was safe and effective in the treatment of chronic allergic dermatitis.

  4. Oxidation of methionine in PrP is dependent upon the oxidant and the amino acid two positions removed

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Background/Introduction. Methionine oxidation has been shown both to be associated with prion formation and implicated in the inhibition of amyloid formation in model systems. This work is based on model systems where hydrogen peroxide was used as an oxidant. Materials and Methods. We developed...

  5. The development and application of a single-cell biosensor for the detection of l-methionine and branched-chain amino acids.

    PubMed

    Mustafi, Nurije; Grünberger, Alexander; Kohlheyer, Dietrich; Bott, Michael; Frunzke, Julia

    2012-07-01

    The detection and quantification of specific metabolites in single bacterial cells is a major goal for industrial biotechnology. We have developed a biosensor based on the transcriptional regulator Lrp that detects intracellular l-methionine and branched-chain amino acids in Corynebacterium glutamicum. In assays, fluorescence output showed a linear relationship with cytoplasmic concentrations of the effector amino acids. In increasing order, the affinity of Lrp for the amino acids is l-valine, l-isoleucine, l-leucine and l-methionine. The sensor was applied for online monitoring and analysis of cell-to-cell variability of l-valine production by the pyruvate dehydrogenase-deficient C. glutamicum strain ΔaceE. Finally, the sensor system was successfully used in a high-throughput (HT) FACS screen for the isolation of amino acid-producing mutants after random mutagenesis of a non-producing wild type strain. These applications illustrate how one of nature's sensor devices - transcriptional regulators - can be used for the analysis, directed evolution and HT screening for microbial strain development.

  6. Inhibition of corneal neovascularization with a nutrient mixture containing lysine, proline, ascorbic acid, and green tea extract.

    PubMed

    Shakiba, Yadollah; Mostafaie, Ali

    2007-10-01

    Corneal neovascularization is a significant, sight-threatening complication of many ocular surface disorders. Various growth factors and proteinases are involved in corneal neovascularization. The data supporting a causal role for vascular endothelial growth factor (VEGF) and matrix metalloproteinases (MMPs) are extensive. Inhibition of VEGF and MMPs is a main strategy for treating corneal neovascularization. Several findings have shown that corneal neovascularization can be reduced by using anti-VEGF and anti-MMPs agents. Efficacy of a nutrient mixture (NM) containing lysine, proline, ascorbic acid, and green tea extract has been demonstrated for reducing VEGF and MMPs secretion by various cells. Moreover, NM can inhibit endothelial cell migration and capillary tube formation. We herein note that topical application of NM is potentially useful for inhibiting corneal neovascularization and restoration of corneal clarity. Further investigations in animal models are needed to place NM alongside corneal neovascularization therapeutics.

  7. Proteomic analysis of different period excretory secretory products from Clonorchis sinensis adult worms: molecular characterization, immunolocalization, and serological reactivity of two excretory secretory antigens-methionine aminopeptidase 2 and acid phosphatase.

    PubMed

    Zheng, Minghui; Hu, Kunhua; Liu, Wei; Li, Hongyu; Chen, Jingfang; Yu, Xinbing

    2013-03-01

    The excretory secretory products (ESP) of Clonorchis sinensis are the causative agents of clonorchiasis and biliary diseases. The parasites' ESP play important roles in host-parasite interactions. The protein compositions of ESP at different secretory times are different and have not been systemically investigated so far. In this study, we collected ESP from six different periods (0-3 h, 3-6 h, 6-12 h, 12-24 h, 24-36 h, and 36-48 h) from C. sinensis adults. Using a shotgun LC-MS/MS analysis, we found 187, 80, 103, 58, 248, and 383 proteins, respectively. Among these proteins, we selected methionine aminopeptidase 2 (MAP-2, presented in 24-36 h and 36-48 h ESP) and acid phosphatase (AP, presented in 3-6 h, 12-24 h, 24-36 h, and 36-48 h ESP) for further study. Bioinformatics analysis showed that CsMAP-2 has metallopeptidase family M24, unique lysine residue-rich and acidic residue-rich domain, SGTS motif, and auto-cleavage point; and that CsAP has possible signal sequence cleavage site, acid phosphate domain, and two histidine acid phosphatases active regions. CsMAP-2 and CsAP's cDNA have 1,425 bp and1,410 bp ORF, encoding 475 and 470 amino acid proteins and weighing 55.3840 kDa and 55.2875 kDa, respectively. MAP-2 and AP were identified as antigens present in the ESP and circulating antigens by immunoblot analysis, which were also found expressing in the eggs, metacercaria, and adult stages of C. sinensis. Immunofluorescence analysis showed that they were located in tegument and intestinal cecum of adult. MTT assay showed that they could inhibit hepatic stellate cell line (LX-2) proliferation. These findings presented the compositions of different period excretory secretary products from C. sinensis adults.

  8. Evaluation of Amino Acids as Turfgrass Nematicides1

    PubMed Central

    Zhang, Yun; Luc, John E.; Crow, William T.

    2010-01-01

    Laboratory experiments revealed that DL-methionine, sodium methionate, potassium methionate, and methionine hydroxyl analog at rates of 224 and 448 kg amino acid/ha reduced the number of Belonolaimus longicaudatus mixed life-stages and Meloidogyne incognita J2 in soil, whereas L-threonine and lysine were not effective in reducing the number of either nematode. Futhermore, greenhouse experiments demonstrated that DL-methionine, sodium methionate, potassium methionate, and methionine hydroxyl analog were equally effective against B. longicaudatus at rates of 112, 224, and 448 kg amino acid/ha, and the highest rate (448 kg amino acid/ha) of all amino acids was more effective in reducing the number of B. longicaudatus than the lower rate. However, phytotoxicity was observed on creeping bentgrass (Agrostis palustris) treated with 448 kg amino acid/ha of methionine hydroxyl analog and DL methionine. In addition, in one of two field experiments on bermudagrass (Cynodon dactylon × C. transvaalensis) turf percentage green cover was increased and the number of B. longicaudatus was reduced by 224 kg amino acid/ha of DL-methionine and potassium methionate compared to untreated controls in one of two trials. PMID:22736861

  9. 47 CRYOPRESERVATION OF BOVINE GERM CELL USING ANTIFREEZE POLYAMINO-ACID (CARBOXYLATED POLY-L-LYSINE).

    PubMed

    Fujikawa, T; Kubota, C; Ando, T; Imamura, S; Tokumaru, M; Yamakuchi, H; Gen, Y; Hyon, S-H

    2016-01-01

    Carboxylated poly-l-lysine (CPLL) is an ampholytic polymer compound, and it is obtained by converting 65% amino groups to carboxyl groups after synthesising ε-poly-l-lysine aqueous solution and succinic anhydride. CPLL has cryoprotective property similar to antifreeze protein, and addition of CPLL into cryopreservation medium improves the post-thaw survival rate of cells and embryos. In this research, we examined the effectiveness of CPLL as a bovine germ cell cryoprotective material. In experiment 1 (in sperm), the conventional cryopreservation medium used for control group was consisted of 6.5% (vol/vol) glycerin, and the cryopreservation medium used for CPLL group was consisted of 3.25% (vol/vol) glycerin and 0.5% CPLL (wt/vol). The post-thaw survival and motility were assessed by using Sperm Motility Analysis System (DITECT Corp., Tokyo, Japan). There was no significant difference for post-thaw survival rate and motility (control v. CPLL; 98.8% v. 96.6% and 69.7% v. 62.2%, respectively). Artificial insemination was carried out in 65 cows (control v. CPLL; 34v. 31), and the conception rate of the CPLL group was higher than that of the control group (80.6% v. 67.6%; P=0.23). In experiment 2 (embryos), the conventional cryopreservation medium used for control group was consisted of 5% (vol/vol) ethylene glycol and 6% (vol/vol) propylene glycol in PBS. In the CPLL group, 7% (wt/vol) CPLL was added to the conventional medium. In vitro fertilization embryos were cryopreserved at Day 7 and Day 8. There was no significant difference in survival rate at 0, 24, and 48h and hatched rate until 72h after thawing (control v. CPLL: 93.6% v. 93.2%, 69.0% v. 64.7%, 56.1% v. 56.3%, 12.9% v. 10.2%, respectively). Embryos obtained by superovulation treatment and in vivo fertilization at Day 7 were cryopreserved using above 2 media, and transferred non-surgically into synchronized recipient cows (1 embryo per animal). Embryo transfer (ET) was carried out in 81 cows (control v

  10. Analysis for availability of amino acid supplements in foods and feeds: biochemical and nutritional implications.

    PubMed

    Ostrowski, H T

    1978-01-01

    In formulated diets based on cereal grains, lysine and/or methionine are usually deficient as well as often being the first amino acids limiting the nutritional value of such diets. Deficiency of these two amino acids in nutritional practice is compensated by synthetic L-lysine and DL-methionine supplementation or by the introduction of various protein sources - rich in lysine and methionine. Among all essential amino acids lysine is most liable and subject to damage during the processing of foods and feeds which can cause the "deepening" of the lysine deficiency not on the total but on the physiologically-available lysine basis. Hence, the simultaneous lysine deficiency and biological "sufficiency" problem is discussed using examples of practical diets in which a balance of biologically-active substances was achieved by the formulation and optimalisation according to the needs of animals, taking into account physiological lysine "accessibility" - "availability". Growth rate, nitrogen balance data and chemical composition of the tissue in long term trials are the most valid indication justifying the quantity of amino acid supplements to the practical diets. Prediction of the practical results of dietary amino acid balance from various short-term chemical and biological tests can give misleading results. Their application in nutritional practice is restricted to particular types of foods/feeds, and to specific processing systems and test conditions. Observations of the appearances of most limiting, dietary amino acids in the blood after the meal do not provide a complete nutritional characteristics of practical rations due to complex regulatory mechanisms in protein and amino acid metabolism much of which are not yet fully understood.

  11. Heterologous Production of Cyanobacterial Mycosporine-Like Amino Acids Mycosporine-Ornithine and Mycosporine-Lysine in Escherichia coli

    PubMed Central

    Katoch, Meenu; Mazmouz, Rabia; Chau, Rocky; Pearson, Leanne A.; Pickford, Russell

    2016-01-01

    ABSTRACT Mycosporine-like amino acids (MAAs) are an important class of secondary metabolites known for their protection against UV radiation and other stress factors. Cyanobacteria produce a variety of MAAs, including shinorine, the active ingredient in many sunscreen creams. Bioinformatic analysis of the genome of the soil-dwelling cyanobacterium Cylindrospermum stagnale PCC 7417 revealed a new gene cluster with homology to MAA synthase from Nostoc punctiforme. This newly identified gene cluster is unusual because it has five biosynthesis genes (mylA to mylE), compared to the four found in other MAA gene clusters. Heterologous expression of mylA to mylE in Escherichia coli resulted in the production of mycosporine-lysine and the novel compound mycosporine-ornithine. To our knowledge, this is the first time these compounds have been heterologously produced in E. coli and structurally characterized via direct spectral guidance. This study offers insight into the diversity, biosynthesis, and structure of cyanobacterial MAAs and highlights their amenability to heterologous production methods. IMPORTANCE Mycosporine-like amino acids (MAAs) are significant from an environmental microbiological perspective as they offer microbes protection against a variety of stress factors, including UV radiation. The heterologous expression of MAAs in E. coli is also significant from a biotechnological perspective as MAAs are the active ingredient in next-generation sunscreens. PMID:27520810

  12. Simultaneous Detection of Dopamine and Uric Acid Using a Poly(l-lysine)/Graphene Oxide Modified Electrode

    PubMed Central

    Zhang, Yuehua; Lei, Wu; Xu, Yujuan; Xia, Xifeng; Hao, Qingli

    2016-01-01

    A novel, simple and selective electrochemical method was investigated for the simultaneous detection of dopamine (DA) and uric acid (UA) on a poly(l-lysine)/graphene oxide (GO) modified glassy carbon electrode (PLL/GO/GCE) by differential pulse voltammetry (DPV). The electrochemically prepared PLL/GO sensory platform toward the oxidation of UA and DA exhibited several advantages, including high effective surface area, more active sites and enhanced electrochemical activity. Compared to the PLL-modified GCE (PLL/GCE), GO-modified GCE and bare GCE, the PLL/GO/GCE exhibited an increase in the anodic potential difference and a remarkable enhancement in the current responses for both UA and DA. For the simultaneous detection of DA and UA, the detection limits of 0.021 and 0.074 μM were obtained, while 0.031 and 0.018 μM were obtained as the detection limits for the selective detection of UA and DA, using DPV in the linear concentration ranges of 0.5 to 20.0 and 0.5 to 35 μM, respectively. In addition, the PLL/GO/GCE demonstrated good reproducibility, long-term stability, excellent selectivity and negligible interference of ascorbic acid (AA). The proposed modified electrode was successfully implemented in the simultaneous detection of DA and UA in human blood serum, urine and dopamine hydrochloride injection with satisfactory results.

  13. Nuclear magnetic resonance studies of amino acids and proteins. Side-chain mobility of methionine in the crystalline amonio acid and in crystallne sperm whale (Physeter catodon) myoglobin

    SciTech Connect

    Keniry, M.A.; Rothgeb, T.M.; Smith, R.L.; Gutowsky, H.S.; Oldfield, E.

    1983-04-12

    Deuterium (/sup 2/H) nuclear magnetic resonance (NMR) spectra and spin-lattice relaxation times (T/sub 1/) were obtained of L-(epsilon-/sup 2/H/sub 3/)methionine, L-(epsilon-/sup 2/H/sub 3/)methionine in a D,L lattice, and (S-methyl-/sup 2/H/sub 3/)methionine in the crystalline solid state, as a function of temperature, in addition to obtaining /sup 2/H T/sub 1/ and line-width results as a function of temperature on (epsilon-/sup 2/H/sub 3/)methionine-labeled sperm whale (Physeter catodon) myoglobins by using the method of magnetic ordering. Also recorded were /sup 13/C cross-polarization ''magic-angle'' sample-spinning NMR spectra of (epsilon-/sup 13/C)methionine-labeled crystalline cyanoferrimyoglobin (at 37.7 MHz, corresponding to a magnetic field strength of 3.52 T) and of the same protein in aqueous solution. (JMT)

  14. Contribution of the net charge to the regulatory effects of amino acids and epsilon-poly(L-lysine) on the gelatinization behavior of potato starch granules.

    PubMed

    Ito, Azusa; Hattori, Makoto; Yoshida, Tadashi; Takahashi, Koji

    2006-01-01

    The effects of lysine (Lys), monosodium glutamate (GluNa), glycine, alanine and epsilon-poly(L-lysine) (PL) with different degrees of polymerization on the gelatinization behavior of potato starch granules were investigated by DSC, viscosity and swelling measurements, microscopic observation, and measurement of the retained amino acid amount to clarify the contribution of the net charge to their regulatory effects on the gelatinization behavior. The amino acids and PL each contributed to an increase in the gelatinization temperature, and a decrease in the peak viscosity and swelling. These effects strongly depended on the absolute value of their net charge. The disappearance of a negative or positive net charge by adjusting the pH value weakened the contribution. The swelling index and size of the potato starch granules changed according to replacement of the swelling medium. The amino acids and PL were easily retained by the swollen potato starch granules according to replacement of the outer solution of the starch granules.

  15. 21 CFR 172.372 - N-Acetyl-L-methionine.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... amino acid methionine formed by addition of an acetyl group to the alpha-amino group of methionine. It... amino acid) by weight of the total protein of the finished food, including the amount naturally present... of the additive contained therein. (2) The amounts of additive and each amino acid contained in...

  16. 21 CFR 172.372 - N-Acetyl-L-methionine.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... Abstracts Service Registry No. 65-82-7) is the derivative of the amino acid methionine formed by addition of... percent L- and DL-methionine (expressed as the free amino acid) by weight of the total protein of the...) The amounts of additive and each amino acid contained in any mixture. (3) Adequate directions for...

  17. 21 CFR 172.372 - N-Acetyl-L-methionine.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... amino acid methionine formed by addition of an acetyl group to the alpha-amino group of methionine. It... amino acid) by weight of the total protein of the finished food, including the amount naturally present... of the additive contained therein. (2) The amounts of additive and each amino acid contained in...

  18. 21 CFR 172.372 - N-Acetyl-L-methionine.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ...-Acetyl-L-methionine (Chemical Abstracts Service Registry No. 65-82-7) is the derivative of the amino acid... provide a total of 3.1 percent L- and DL-methionine (expressed as the free amino acid) by weight of the... contained therein. (2) The amounts of additive and each amino acid contained in any mixture. (3)...

  19. 21 CFR 172.372 - N-Acetyl-L-methionine.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... amino acid methionine formed by addition of an acetyl group to the alpha-amino group of methionine. It... amino acid) by weight of the total protein of the finished food, including the amount naturally present... of the additive contained therein. (2) The amounts of additive and each amino acid contained in...

  20. Methionine metabolism in Yucatan miniature swine.

    PubMed

    McBreairty, Laura E

    2016-06-01

    Methionine is an essential amino acid which when not incorporated into protein, can be converted to S-adenosylmethionine, the universal methyl donor in over 200 transmethylation reactions, which include creatine and phosphatidylcholine (PC) synthesis, as well as deoxyribonucleic acid (DNA) methylation. Following transmethylation, homocysteine is formed, which can be converted to cysteine via transsulfuration or remethylated to methionine by receiving a methyl group from folate or betaine. Changes to methyl group availability in utero can lead to permanent changes in epigenetic patterns of DNA methylation, which has been implicated in "fetal programming", a phenomenon associated with poor nutrition during fetal development that results in low birth weight and disease in later life. It has been shown that programming can also occur in the neonate. Our global objective was to understand how the variability of nutrients involved in methionine metabolism can affect methionine and methyl group availability. We hypothesize that nutrients that converge on methionine metabolism can affect methionine availability for its various functions. In this thesis, we used intrauterine growth restricted (IUGR) piglets to investigate whether a global nutritional insult in utero can lead to a perturbed methionine metabolism. Our results demonstrate that IUGR piglets have a lower capacity to dispose of homocysteine via both transsulfuration and remethylation pathways, as well as a lower incorporation of methyl groups into PC. The second objective of this thesis was to determine whether variation in methionine supply and demand can affect methionine availability. We demonstrated that stimulating either acute or chronic creatine synthesis leads to lower methyl incorporation into protein and PC in pigs. Furthermore, when methionine is limiting, supplementation with either folate or betaine leads to higher methionine availability for protein synthesis. Finally, because creatine is

  1. Factors influencing methionine toxicity in young bobwhite quail

    USGS Publications Warehouse

    Serafin, J.A.

    1981-01-01

    Young Bobwhite quail (Colinus virginianus) were fed low and adequate protein purified diets with and without excess methionine to evaluate factors affecting methionine toxicity. Growth of quail fed an adequate protein (27%) diet, without supplemental glycine, was depressed by 1.75% and 2.25% excess methionine. Supplemental glycine (.3%) alleviated growth depression caused by 2.25% excess methionine. Quail fed 1.75% and 2.25% excess methionine developed signs of toxicity characterized by weakness, a lowered, outstretched neck when moving, and ataxia. In addition, quail would fall on their sides when disturbed and spin with their heads retracted. These conditions were transient in nature. Growth of quail fed a low protein (18.9%) diet was depressed by 1% and 1.5% excess methionine and DL-homocystine. Quail fed 1% and 1.5% excess methionine in this diet also developed signs of toxicity, the incidence of which was greater and the duration longer than occurred with quail fed adequate protein. Supplementing a low protein (20.15%) diet with .3% or .6% glycine or threonine or a combination of these amino acids did not alleviate growth depression caused by 1.5% excess methionine; however, 2% and 3% supplemental glycine were somewhat effective. Supplements of glycine (2%, 3%) and threonine (1%) completely reversed growth depression from 1% excess methionine but did not influence growth of controls, indicating that both amino acids counteract methionine toxicity. Both glycine and threonine alone improved growth by about the same extent in diets with 1% or 1.5% excess methionine; however, these amino acids alleviated less than 30% of the growth depression resulting from 1.5% excess methionine. The effectiveness of glycine in alleviating methionine toxicity in a low protein diet was decreased, and hemoglobin levels were depressed with 1.5% excess methionine compared to less amounts.

  2. Heterologous Production of Cyanobacterial Mycosporine-Like Amino Acids Mycosporine-Ornithine and Mycosporine-Lysine in Escherichia coli.

    PubMed

    Katoch, Meenu; Mazmouz, Rabia; Chau, Rocky; Pearson, Leanne A; Pickford, Russell; Neilan, Brett A

    2016-10-15

    Mycosporine-like amino acids (MAAs) are an important class of secondary metabolites known for their protection against UV radiation and other stress factors. Cyanobacteria produce a variety of MAAs, including shinorine, the active ingredient in many sunscreen creams. Bioinformatic analysis of the genome of the soil-dwelling cyanobacterium Cylindrospermum stagnale PCC 7417 revealed a new gene cluster with homology to MAA synthase from Nostoc punctiforme This newly identified gene cluster is unusual because it has five biosynthesis genes (mylA to mylE), compared to the four found in other MAA gene clusters. Heterologous expression of mylA to mylE in Escherichia coli resulted in the production of mycosporine-lysine and the novel compound mycosporine-ornithine. To our knowledge, this is the first time these compounds have been heterologously produced in E. coli and structurally characterized via direct spectral guidance. This study offers insight into the diversity, biosynthesis, and structure of cyanobacterial MAAs and highlights their amenability to heterologous production methods.

  3. Acetylation dictates the morphology of nanophase biosilica precipitated by a 14-amino acid leucine-lysine peptide.

    PubMed

    Lutz, Helmut; Jaeger, Vance; Bonn, Mischa; Pfaendtner, Jim; Weidner, Tobias

    2017-02-01

    N-terminal acetylation is a commonly used modification technique for synthetic peptides, mostly applied for reasons of enhanced stability, and in many cases regarded as inconsequential. In engineered biosilification - the controlled deposition of silica for nanotechnology applications by designed peptides - charged groups often play a deciding role. Here we report that changing the charge by acetylation of a 14-amino acid leucine-lysine (LK) peptide dramatically changes the morphology of precipitated biosilica; acetylated LK peptides produce nano-spheres, whereas nano-wires are precipitated by the same peptide in a non-acetylated form. By using interface-specific vibrational spectroscopy and coarse-grained molecular simulations, we show that this change in morphology is not the result of modified peptide-silica interactions, but rather caused by the stabilization of the hydrophobic core of peptide aggregates created by the removal of a peptide charge upon acetylation. These results should raise awareness of the potential impact of N-terminal modifications in peptide applications. Copyright © 2016 European Peptide Society and John Wiley & Sons, Ltd.

  4. Cyclen Grafted with poly[(Aspartic acid)-co-Lysine]: Preparation, Assembly with Plasmid DNA, and in Vitro Transfection Studies.

    PubMed

    Ma, Chunying; Zhang, Jin; Guo, Liwen; Du, Changguo; Song, Ping; Zhao, Baojing; Li, Ling; Li, Chao; Qiao, Renzhong

    2016-01-04

    Development of safe and effective gene carriers is the key to the success of gene therapy. Nowadays, it is still required to develop new methods to improve nonviral gene delivery efficiency. Herein, copolymers of poly[(aspartic acid)-co-lysine] grafted with cyclen (cyclen-pAL) were designed and evaluated for efficient gene delivery. Two copolymers with different Asp/Lys block ratios were prepared and characterized by NMR and gel permeation chromatography analysis. Agarose gel retardation, circular dichroism, and fluorescent quenching assays showed the strong DNA-binding and protection ability for the title compounds. Atomic force microscopy studies clearly delineated uniform DNA globules with a diameter around 100 nm, induced by cyclen-pAL. By grafting cyclen on Asp, relatively high gene delivery efficiency and low cytotoxicity of the modified copolymers were achieved compared with their parent compounds. The present work might help to develop strategies for design and modification of polypeptide copolymers, which may also be applied to favorable gene expression and delivery.

  5. Isoxazole‐Derived Amino Acids are Bromodomain‐Binding Acetyl‐Lysine Mimics: Incorporation into Histone H4 Peptides and Histone H3

    PubMed Central

    Sekirnik (née Measures), Angelina R.; Hewings, David S.; Theodoulou, Natalie H.; Jursins, Lukass; Lewendon, Katie R.; Jennings, Laura E.; Rooney, Timothy P. C.; Heightman, Tom D.

    2016-01-01

    Abstract A range of isoxazole‐containing amino acids was synthesized that displaced acetyl‐lysine‐containing peptides from the BAZ2A, BRD4(1), and BRD9 bromodomains. Three of these amino acids were incorporated into a histone H4‐mimicking peptide and their affinity for BRD4(1) was assessed. Affinities of the isoxazole‐containing peptides are comparable to those of a hyperacetylated histone H4‐mimicking cognate peptide, and demonstrated a dependence on the position at which the unnatural residue was incorporated. An isoxazole‐based alkylating agent was developed to selectively alkylate cysteine residues in situ. Selective monoalkylation of a histone H4‐mimicking peptide, containing a lysine to cysteine residue substitution (K12C), resulted in acetyl‐lysine mimic incorporation, with high affinity for the BRD4 bromodomain. The same technology was used to alkylate a K18C mutant of histone H3. PMID:27264992

  6. Methionine restriction and lifespan control

    PubMed Central

    Lee, Byung Cheon; Kaya, Alaattin; Gladyshev, Vadim N.

    2016-01-01

    Dietary restriction (DR) without malnutrition is associated with longevity in various organisms. However, it has also been shown that reduced calorie intake is often ineffective in extending lifespan. Selecting optimal dietary regimens for DR studies is complicated, as the same regimen may lead to different outcomes depending on genotype and environmental factors. Recent studies suggested that interventions such as moderate protein restriction with/without adequate nutrition (e.g. particular amino acids or carbohydrates) may have additional beneficial effects mediated by certain metabolic and hormonal factors implicated in the biology of aging, regardless of total calorie intake. In particular, it was shown that restriction of a single amino acid, methionine, can mimic the effects of DR and extend lifespan in various model organisms. We discuss beneficial effects of methionine-restricted (MR) diet, the molecular pathways involved, and the use of this regimen in longevity interventions. PMID:26663138

  7. Effect of l-lysine-assisted surface grafting for nano-hydroxyapatite on mechanical properties and in vitro bioactivity of poly(lactic acid-co-glycolic acid).

    PubMed

    Liuyun, Jiang; Lixin, Jiang; Chengdong, Xiong; Lijuan, Xu; Ye, Li

    2016-01-01

    It is promising and challenging to study surface modification for nano-hydroxyapatite to improve the dispersion and enhance the mechanical properties and bioactivity of poly(lactic acid-co-glycolic acid). In this paper, we designed an effective new surface grafting with the assist of l-lysine for nano-hydroxyapatite, and the nano-hydroxyapatite surface grafted with the assist of l-lysine (g-nano-hydroxyapatite) was incorporated into poly(lactic acid-co-glycolic acid) to develop a series of g-nano-hydroxyapatite/poly(lactic acid-co-glycolic acid) nano-composites. The surface modification reaction for nano-hydroxyapatite, the mechanical properties, and in vitro human osteoblast-like cell (MG-63) response were characterized and investigated by Fourier transformation infrared, thermal gravimetric analysis, dispersion test, electromechanical universal tester, differential scanning calorimeter measurements, and in vitro cells culture experiment. The results showed that the grafting amount on the surface of nano-hydroxyapatite was enhanced with the increase of l-lysine, and the dispersion of nano-hydroxyapatite was improved more, so that it brought about better promotion crystallization and more excellent mechanical enhancement effect for poly(lactic acid-co-glycolic acid), comparing with the unmodified nano-hydroxyapatite. Moreover, the cells' attachment and proliferation results confirmed that the incorporation of the g-nano-hydroxyapatite into poly(lactic acid-co-glycolic acid) exhibited better biocompatibility than poly(lactic acid-co-glycolic acid). The above results indicated that the new surface grafting with the assist of l-lysine for nano-hydroxyapatite was an ideal novel surface modification method, which brought about better mechanical enhancement effect and in vitro bioactivity for poly(lactic acid-co-glycolic acid) with adding higher g-nano-hydroxyapatite content, suggesting it had a great potential to be used as bone fracture internal fixation materials

  8. [Effect of amino acid supplements to barley meal on the nitrogen metabolism of growing castrated male swine (20-65 kg live weight)].

    PubMed

    Wecke, C; Gebhardt, G

    1981-03-01

    In 56 N-balance experiments of the influence of differentiated amino acid supplements to coarse barley meal enriched with energy, minerals and additives on the nitrogen metabolism of castrated male pigs, was investigated. The joint supplement of lysine and methionine remained without result in comparison with the sole supplementation of lysine. Only the additional supplementation of threonine resulted in the further improvement of protein utilisation. The results corroborate the effect of the amino acid lysine limiting the performance in barley protein and prove that threonine takes the second place in the sequence of limitation.

  9. Local flexibility facilitates oxidization of buried methionine residues.

    PubMed

    Xu, Kuiran; Uversky, Vladimir N; Xue, Bin

    2012-06-01

    In proteins, all amino acid residues are susceptible to oxidation by various reactive oxygen species (ROS), with methionine and cysteine residues being particularly sensitive to oxidation. Methionine oxidation is known to lead to destabilization and inactivation of proteins, and oxidatively modified proteins can accumulate during aging, oxidative stress, and in various age-related diseases. Although the efficiency of a given methionine oxidation can depend on its solvent accessibility (evaluated from a protein structure as the accessible surface area of the corresponding methionine residue), many experimental results on oxidation rate and oxidation sites cannot be unequivocally explained by the methionine solvent accessible surface area alone. In order to explore other possible mechanisms, we analyzed a set of seventy-one oxidized methionines contained in thirty-one proteins by various bioinformatics tools. In which, 41% of the methionines are exposed, 15% are buried but with various degree of flexibility, and the rest 44% are buried and structured. Buried but highly flexible methionines can be oxidized. Buried and less flexible methionines can acquire additional local structural flexibility from flanking regions to facilitate the oxidation. Oxidation of buried and structured methionine can also be promoted by the oxidation of neighboring methionine that is more exposed and/or flexible. Our data are consistent with the hypothesis that protein structural flexibility represents another important factor favoring the oxidation process.

  10. Limiting amino acids in bengal gram (Cicer arietinum) as determined from blood amino acid levels and amino acid supplementation studies in the rat.

    PubMed

    Khader, V; Rao, S V

    1982-01-01

    The limiting amino acids of Bengal gram (Cicer arietinum) were determined from plasma amino acid score and ratio and growth response of weanling rats to supplements of amino acids. The results indicated that methionine, threonine and tryptophan are the most limiting amino acids. Protein efficiency ratio of raw and cooked Bengal gram fed at a dietary level of 10% protein increased from 2.7 to 3.7 and 2.4 to 3.4, respectively, on supplementing the diets with methionine, threonine and tryptophan. Plasma levels of lysine, methionine, threonine and tryptophan were similar in rats fed raw or cooked Bengal gram, indicating that the trypsin or other inhibitors that may be present in the raw gram do not affect the biological availability of these amino acids.

  11. Quantum Computational Calculations of the Ionization Energies of Acidic and Basic Amino Acids: Aspartate, Glutamate, Arginine, Lysine, and Histidine

    NASA Astrophysics Data System (ADS)

    de Guzman, C. P.; Andrianarijaona, M.; Lee, Y. S.; Andrianarijaona, V.

    An extensive knowledge of the ionization energies of amino acids can provide vital information on protein sequencing, structure, and function. Acidic and basic amino acids are unique because they have three ionizable groups: the C-terminus, the N-terminus, and the side chain. The effects of multiple ionizable groups can be seen in how Aspartate's ionizable side chain heavily influences its preferred conformation (J Phys Chem A. 2011 April 7; 115(13): 2900-2912). Theoretical and experimental data on the ionization energies of many of these molecules is sparse. Considering each atom of the amino acid as a potential departing site for the electron gives insight on how the three ionizable groups affect the ionization process of the molecule and the dynamic coupling between the vibrational modes. In the following study, we optimized the structure of each acidic and basic amino acid then exported the three dimensional coordinates of the amino acids. We used ORCA to calculate single point energies for a region near the optimized coordinates and systematically went through the x, y, and z coordinates of each atom in the neutral and ionized forms of the amino acid. With the calculations, we were able to graph energy potential curves to better understand the quantum dynamic properties of the amino acids. The authors thank Pacific Union College Student Association for providing funds.

  12. Characterization of isolated yeast growth response to methionine analogs.

    PubMed

    Saengkerdsub, Suwat; Lingbeck, Jody M; Wilkinson, Heather H; O'Bryan, Corliss A; Crandall, Philip G; Muthaiyan, Arunachalam; Biswas, Debabrata; Ricke, Steven C

    2013-01-01

    Methionine is one of the first limiting amino acids in poultry nutrition. The use of methionine-rich natural feed ingredients, such as soybean meal or rapeseed meal may lead to negative environmental consequences. Amino acid supplementation leads to reduced use of protein-rich ingredients. The objectives of this study were isolation of potentially high content methionine-containing yeasts, quantification of methionine content in yeasts and their respective growth response to methionine analogs. Minimal medium was used as the selection medium and the isolation medium of methionine-producing yeasts from yeast collection and environmental samples, respectively. Two yeasts previously collected along with six additional strains isolated from Caucasian kefir grains, air-trapped, cantaloupe, and three soil samples could grow on minimal medium. Only two of the newly isolated strains, K1 and C1, grew in minimal medium supplied with either methionine analogs ethionine or norleucine at 0.5% (w/v). Based on large subunit rRNA sequences, these isolated strains were identified as Pichia udriavzevii/Issatchenkia orientalis. P. kudriavzevii/I. orentalis is a generally recognized as a safe organism. In addition, methionine produced by K1 and C1 yeast hydrolysate yielded 1.3 ± 0.01 and 1.1 ± 0.01 mg g(-1) dry cell. Yeast strain K1 may be suitable as a potential source of methionine for dietary supplements in organic poultry feed but may require growth conditions to further increase their methionine content.

  13. Oral delivery of zoledronic acid by non-covalent conjugation with lysine-deoxycholic acid: In vitro characterization and in vivo anti-osteoporotic efficacy in ovariectomized rats.

    PubMed

    Jeon, Ok-Cheol; Seo, Dong-Hyun; Kim, Han-Sung; Byun, Youngro; Park, Jin Woo

    2016-01-20

    We assessed the possibility of changing the route of administration of zoledronic acid to an oral dosage form and its therapeutic efficacy in an estrogen-deficient osteoporosis rat model. To enhance oral bioavailability, we formed an ionic complex by electrostatic conjugation of zoledronic acid with lysine-linked deoxycholic acid (Lys-DOCA, an oral absorption enhancer). After forming the complex, the characteristic crystalline features of pure zoledronic acid disappeared completely in the powder X-ray diffractogram and differential scanning calorimetry thermogram, indicating that zoledronic acid existed in an amorphous form in the complex. In vitro permeabilities of zoledronic acid/Lys-DOCA (1:1) (ZD1) and zoledronic acid/Lys-DOCA (1:2) (ZD2) complex across Caco-2 cell monolayers were 2.47- and 4.74-fold higher than that of zoledronic acid, respectively. Upon intra-jejunal administration to rats, the intestinal absorption of zoledronic acid was increased significantly and the resulting oral bioavailability of the ZD2 complex was determined to be 6.76±2.59% (0.548±0.161% for zoledronic acid). Ovariectomized (OVX) rats showed 122% increased bone mineral density versus the OVX control at 12weeks after treatment with once weekly oral administration of ZD2 complex (16μg/kg of zoledronic acid). Furthermore, rats treated with ZD2 complex orally showed significant improvement in the parameters of trabecular microarchitecture and bone strength: 149% higher bone volume fraction (BV/TV), 115% higher trabecular number (Tb.N), and 56% higher mean maximum load (Fmax) than in the OVX group. The trabecular microstructure and bone mechanical properties in the oral zoledronic acid group were not significantly changed compared with the OVX control. Thus, the oral ZD2 complex inhibited osteoporosis progression effectively by promoting osteogenesis and trabecular connectivity. The oral ZD2 complex would be expected to improve patient compliance by replacing the conventional

  14. Metabolism of 5-methylthioribose to methionine

    SciTech Connect

    Miyazaki, J.H.; Yang, S.F.

    1987-06-01

    During ethylene biosynthesis, the H/sub 3/CS-group of S-adenosylmethionine is released as 5'-methylthioadenosine, which is recycled to methionine via 5-methylthioribose (MTR). In mungbean hypocotyls and cell-free extracts of avocado, (/sup 14/C)MTR was converted into labeled methionine via 2-keto-4-methylthiobutyric acid (KMB) and 2-hydroxy-4-methylthiobutyric acid (HMB), as intermediates. Incubation of (ribose-U-/sup 14/C)MTR with avocado extract resulted in the production of (/sup 14/C)formate, indicating the conversion of MTR to KMB involves a loss of formate, presumably from C-1 of MTR. Tracer studies showed that KMB was converted readily in vivo and in vitro to methionine, while HMB was converted much more slowly. The conversion of KMB to methionine by dialyzed avocado extract requires an amino donor. Among several potential donors examined, L-glutamine was the most efficient. Anaerobiosis inhibited only partially the oxidation of MTR to formate, KMB/HMB, and methionine by avocado extract. The role of O/sub 2/ in the conversion of MTR to methionine is discussed.

  15. The regulatory effect of citric acid on the co-production of poly(ε-lysine) and poly(L-diaminopropionic acid) in Streptomyces albulus PD-1.

    PubMed

    Xia, Jun; Xu, Zhaoxian; Xu, Hong; Feng, Xiaohai; Bo, Fangfang

    2014-10-01

    Streptomyces albulus PD-1 can co-produce antimicrobial homo-polymers poly(ε-lysine) (ε-PL) and poly(L-diaminopropionic acid) (PDAP). In this study, a novel feeding strategy of citric acid coupled with glucose-(NH4)2SO4 feeding was employed to S. albulus PD-1. When the pH of the culture broth dropped to 4.0, the feeding solution was added continuously to maintain the concentrations of glucose and citric acid at 10 and 4 g L(-1), respectively. As a result, the final concentration of ε-PL increased from 21.7 to 29.7 g L(-1) and the final concentration of PDAP decreased from 4.8 to 3.2 g L(-1). Assays on intracellular nucleotide levels and key enzyme activities were performed to elucidate the underlying regulation mechanism. The addition of citric acid increased NADH/NAD(+) ratio and decreased intracellular ATP level; meanwhile, the activities of pyruvate kinase, citrate synthase and isocitrate dehydrogenase decreased while aspartate aminotransferase activity increased. Therefore, we deduced that citric acid feeding resulted in metabolic flux redistribution at the node of phosphoenolpyruvate; the metabolic pathway from phosphoenolpyruvate directed into tricarboxylic acid cycle was weakened and thus PDAP production was inhibited. On the other hand, the metabolic pathway from phosphoenolpyruvate directed into oxaloacetate and L-aspartate was enhanced, thereby improving ε-PL production. This fermentation strategy may be potentially useful in ε-PL production because it can effectively inhibit the formation of by-products, such as PDAP.

  16. Efficacy of a Complex of 5-Aminolevulinic Acid and Glycyl-Histidyl-Lysine Peptide on Hair Growth

    PubMed Central

    Sim, Hyun Bo; Jang, Yong Hyun; Lee, Seok-Jong; Kim, Do Won; Yim, Soon-Ho

    2016-01-01

    Background Pattern hair loss is a very common problem. Although effective therapeutics for the treatment of pattern hair loss have been used, novel therapeutic modalities are still required to enhance hair growth. Objective We investigated the efficacy and safety of a complex (ALAVAX) of 5-aminolevulinic acid (5-ALA) and glycyl-histidyl-lysine (GHK) peptide for the treatment of pattern hair loss. Methods Forty-five patients with male pattern hair loss were treated with ALAVAX 100 mg/ml (group A), ALAVAX 50 mg/ml (group B) or placebo (group C) once a day for 6 months. Total hair count, hair length, hair thickness, patient's assessment and adverse events were evaluated at month 1, 3, and 6. Results An increase in hair count for 6 months was 52.6 (p<0.05) in group A, 71.5 (p<0.05) in group B, and 9.6 in group C. The ratio of changes in hair count between group B (2.38) and group C (1.21) at 6 months showed a statistically significant difference (p<0.05). The proportion above good satisfaction was higher in group A (26.7%) than in the other groups (group B: 14.3%, group C: 7.1%). There was no statistically significant difference in hair length and hair thickness among 3 groups at 6 months. There was no adverse event in 3 groups. Conclusion Our study showed that a complex of 5-ALA and GHK peptide may be considered as one of the complementary agents for the treatment of male pattern hair loss. PMID:27489425

  17. Synthesis of Met-enkephalin by solution-phase peptide synthesis methodology utilizing para-toluene sulfonic acid as N-terminal masking of l-methionine amino acid.

    PubMed

    Khan, Riaz A

    2016-12-01

    The Met-enkephalin, Tyr-Gly-Gly-Phe-Met, was synthesized by the solution-phase synthesis (SPS) methodology employing -OBzl group as carboxyls' protection, while the t-Boc groups were employed for the N-terminal α-amines' protection for the majority of the amino acids of the pentapeptide sequence. The l-methionine (l-Met) amino acid was used as PTSA.Met-OBzl obtained from the simultaneous protection of the α-amino, and carboxyl group with para-toluene sulfonic acid (PTSA) and as-OBzl ester, respectively in a C-terminal start of the 2 + 2 + 1 fragments condensation convergent synthetic approach. The protection strategy provided a short, single-step, simultaneous, orthogonal, nearly quantitative, robust, and stable process to carry through the protected l-methionine and l-phenylalanine coupling without any structural deformities during coupling and workups. The structurally confirmed final pentapeptide product was feasibly obtained in good yields through the current approach.

  18. Current topics in the biotechnological production of essential amino acids, functional amino acids, and dipeptides.

    PubMed

    Mitsuhashi, Satoshi

    2014-04-01

    Amino acids play important roles in both human and animal nutrition and in the maintenance of health. Here, amino acids are classified into three groups: first, essential amino acids, which are essential to nutrition; second, functional amino acids, recently found to be important in the promotion of physiological functions; and third, dipeptides, which are used to resolve problematic features of specific free amino acids, such as their instability or insolubility. This review focusses on recent researches concerning the microbial production of essential amino acids (lysine and methionine), functional amino acids (histidine and ornithine), and a dipeptide (L-alanyl-L-glutamine).

  19. Production of the amino acids l-glutamate, l-lysine, l-ornithine and l-arginine from arabinose by recombinant Corynebacterium glutamicum.

    PubMed

    Schneider, Jens; Niermann, Karin; Wendisch, Volker F

    2011-07-10

    Amino acid production processes with Corynebacterium glutamicum are based on media containing glucose from starch hydrolysis or fructose and sucrose as present in molasses. Simultaneous utilization of various carbon sources, including glucose, fructose and sucrose, in blends is a typical characteristic of this bacterium. The renewable non-food carbon source arabinose, which is present in hemicellulosic hydrolysates, cannot be utilized by most C. glutamicum strains. Heterologous expression of the araBAD operon from Escherichia coli in the wild-type and in an l-lysine producing strain of C. glutamicum was shown to enable production of l-glutamate and l-lysine, respectively, from arabinose as sole carbon source. l-Ornithine and l-arginine producing strains were constructed and shown to produce l-ornithine and l-arginine from arabinose when araBAD from E. coli was expressed. Moreover, the recombinant strains produced l-glutamate, l-lysine, l-ornithine and l-arginine respectively, from arabinose also when glucose-arabinose blends were used as carbon sources.

  20. Proteomic and Biochemical Studies of Lysine Malonylation Suggest Its Malonic Aciduria-associated Regulatory Role in Mitochondrial Function and Fatty Acid Oxidation*

    PubMed Central

    Colak, Gozde; Pougovkina, Olga; Dai, Lunzhi; Tan, Minjia; te Brinke, Heleen; Huang, He; Cheng, Zhongyi; Park, Jeongsoon; Wan, Xuelian; Liu, Xiaojing; Yue, Wyatt W.; Wanders, Ronald J. A.; Locasale, Jason W.; Lombard, David B.; de Boer, Vincent C. J.; Zhao, Yingming

    2015-01-01

    The protein substrates of sirtuin 5-regulated lysine malonylation (Kmal) remain unknown, hindering its functional analysis. In this study, we carried out proteomic screening, which identified 4042 Kmal sites on 1426 proteins in mouse liver and 4943 Kmal sites on 1822 proteins in human fibroblasts. Increased malonyl-CoA levels in malonyl-CoA decarboxylase (MCD)-deficient cells induces Kmal levels in substrate proteins. We identified 461 Kmal sites showing more than a 2-fold increase in response to MCD deficiency as well as 1452 Kmal sites detected only in MCD−/− fibroblast but not MCD+/+ cells, suggesting a pathogenic role of Kmal in MCD deficiency. Cells with increased lysine malonylation displayed impaired mitochondrial function and fatty acid oxidation, suggesting that lysine malonylation plays a role in pathophysiology of malonic aciduria. Our study establishes an association between Kmal and a genetic disease and offers a rich resource for elucidating the contribution of the Kmal pathway and malonyl-CoA to cellular physiology and human diseases. PMID:26320211

  1. Bio-inspired nitrile hydration by peptidic ligands based on L-cysteine, L-methionine or L-penicillamine and pyridine-2,6-dicarboxylic acid.

    PubMed

    Byrne, Cillian; Houlihan, Kate M; Devi, Prarthana; Jensen, Paul; Rutledge, Peter J

    2014-12-12

    Nitrile hydratase (NHase, EC 4.2.1.84) is a metalloenzyme which catalyses the conversion of nitriles to amides. The high efficiency and broad substrate range of NHase have led to the successful application of this enzyme as a biocatalyst in the industrial syntheses of acrylamide and nicotinamide and in the bioremediation of nitrile waste. Crystal structures of both cobalt(III)- and iron(III)-dependent NHases reveal an unusual metal binding motif made up from six sequential amino acids and comprising two amide nitrogens from the peptide backbone and three cysteine-derived sulfur ligands, each at a different oxidation state (thiolate, sulfenate and sulfinate). Based on the active site geometry revealed by these crystal structures, we have designed a series of small-molecule ligands which integrate essential features of the NHase metal binding motif into a readily accessible peptide environment. We report the synthesis of ligands based on a pyridine-2,6-dicarboxylic acid scaffold and L-cysteine, L-S-methylcysteine, L-methionine or L-penicillamine. These ligands have been combined with cobalt(III) and iron(III) and tested as catalysts for biomimetic nitrile hydration. The highest levels of activity are observed with the L-penicillamine ligand which, in combination with cobalt(III), converts acetonitrile to acetamide at 1.25 turnovers and benzonitrile to benzamide at 1.20 turnovers.

  2. Linkages in thermal copolymers of lysine

    NASA Technical Reports Server (NTRS)

    Fox, S. W.; Suzuki, F.

    1976-01-01

    The thermal copolymerization of lysine with other alpha-amino acids has been studied further. The identity of the second amino acid influences various properties of the polymer obtained, including the proportion of alpha and epsilon linkages of lysine. A review of linkages in proteinoids indicates alpha and beta linkages for aspartic acid, alpha and gamma linkages for glutamic acid, alpha and epsilon linkages for lysine, and alpha linkages for other amino acids. Thermal proteinoids are thus more complex in types of linkage than are proteins

  3. Linkages in thermal copolymers of lysine

    NASA Technical Reports Server (NTRS)

    Fox, S. W.; Suzuki, F.

    1975-01-01

    The thermal copolymerization of lysine with other alpha-amino acids was studied. The identity of the second amino acid influences various properties of the polymer obtained, including the proportion of alpha and epsilon linkages of lysine. A review of linkages in proteinoids indicates alpha and beta linkages for aspartic acid, alpha and gamma linkages for glutamic acid, alpha and epsilon linkages for lysine, and alpha linkages for other amino acids. Thermal proteinoids are thus more complex in types of linkage than are proteins.

  4. Electrodeposited reduced graphene oxide incorporating polymerization of l-lysine on electrode surface and its application in simultaneous electrochemical determination of ascorbic acid, dopamine and uric acid.

    PubMed

    Zhang, Dongdong; Li, Lingzhi; Ma, Weina; Chen, Xia; Zhang, Yanmin

    2017-01-01

    This paper demonstrates a novel strategy for the construction of a graphene hybrid composites film, which was fabricated by electrodeposited reduced graphene oxide (ERGO) incorporating polymerization of l-lysine (PLL) onto glassy carbon electrode (GCE). Here we show that graphene films can be prepared on electrodes directly from GO dispersions by one-step electrodeposition technique based on electropolymerized PLL as a positively charged polymer interface to adsorb negatively charged GO nanosheets through electrostatic attraction. The thickness of graphene film can be easily controlled by using the electrodeposition technique, a distinct advantage over previously developed methods. The electrochemically reduced process of GO and electropolymerization of l-lysine were investigated by cyclic voltammetry with a wide potential range. The surface morphology of the modified electrode was characterized by scanning electron microscopy. The ERGO/PLL/GCE shows conducive to electron transfer kinetics for Fe(CN)6(3-)/Fe(CN)6(4-) redox probes, compared with bare GCE, PLL/GCE and ERGO/GCE. The electrochemical behaviors of ascorbic acid (AA), dopamine (DA) and uric acid (UA) at ERGO/PLL/GCE were investigated by cyclic voltammetry, and the results suggest that the modified electrode exhibits enhanced electrocatalytic activity toward these important molecules. Under physiological condition and in the co-existence system of AA, DA and UA, the ERGO/PLL/GCE showed linear voltammetric responses in the concentration of 100μM-1200μM for AA, 2.0μM-60μM for DA and 20μM-200μM for UA, and with the detection limits (S/N=3) of 2.0μM, 0.10μM and 0.15μM for AA, DA and UA, respectively. The developed method has been applied to simultaneous determination of AA, DA and UA in human urine with satisfactory recoveries of 104.2%, 95.4% and 99.9%, respectively. This work demonstrates that the attractive features of ERGO/PLL provide promising applications in simultaneous determination of AA, DA

  5. The same substitution, glutamic acid----lysine at position 501, occurs in three alloalbumins of Asiatic origin: albumins Vancouver, Birmingham, and Adana.

    PubMed Central

    Huss, K; Madison, J; Ishioka, N; Takahashi, N; Arai, K; Putnam, F W

    1988-01-01

    A strategy is described for identifying structural changes in genetic variants of human serum albumin (alloalbumins). By use of this strategy we have determined an amino acid substitution in three alloalbumins of Asiatic origin. The same amino acid exchange, glutamic acid----lysine at position 501, occurs in albumins Vancouver and Birmingham, both from families that migrated from northern India, and also in albumin Adana from Turkey. This exchange corresponds to a single base mutation in the codon GAG to AAG and accords with the slow mobility of the three albumins at pH 8.6. Each of the three alloalbumins had been reported to be a new variant, yet they have the same substitution. These results emphasize the need for structural study of genetic variants that have been differentiated only by nonspecific physical criteria such as dye binding and electrophoretic mobility. We know of no other description of the substitution involved in an alloalbumin originating from the Indian subcontinent. However, the same change of glutamic acid----lysine at position 501 may be present in several other named variants reported for populations in north India and the surrounding regions. Images PMID:2901102

  6. Influence of methionine supply on the response of lactational performance of dairy cows to supplementary folic acid and vitamin B12.

    PubMed

    Preynat, A; Lapierre, H; Thivierge, M C; Palin, M F; Matte, J J; Desrochers, A; Girard, C L

    2009-04-01

    The present experiment was undertaken to determine if the effects of supplementary folic acid on lactational performance were caused by improved methylneogenesis and if the supply in vitamin B(12) could affect this metabolic pathway. In this eventuality, supplementary Met, a major source of preformed methyl groups, should reduce the requirements for these vitamins. Sixty multiparous Holstein cows were assigned to 10 blocks of 6 cows each according to their previous milk production. Within each block, 3 cows were fed a diet estimated to supply Met as 1.83% metabolizable protein and 3 cows were fed the same diet supplemented with 18 g of rumen-protected methionine (RPM) to supply Met as 2.23% of metabolizable protein. Within each level of Met, cows received no vitamin supplement or weekly intramuscular injections of 160 mg of folic acid alone or combined with 10 mg of vitamin B(12) from 3 wk before to 16 wk after calving. There was no treatment effect on dry matter intake during pre- and postcalving periods: 13.4 +/- 0.4 and 21.8 +/- 0.4 kg/d, respectively. Milk production was not affected by RPM supplementation. Folic acid and vitamin B(12) given together tended to increase milk production during the 16 wk of lactation. This effect was more pronounced during the first 4 wk of lactation: 37.5, 37.7, and 40.3 +/- 0.9 kg/d for cows receiving no vitamin supplement, folic acid alone, or folic acid combined with vitamin B(12), respectively. Milk fat yield was not affected by treatments. Lactose, crude protein, and total solid yields were greater, in early lactation, in cows injected with folic acid and vitamin B(12) together but this effect diminished as lactation progressed. Intramuscular injections of folic acid alone or combined with vitamin B(12) tended to decrease plasma concentrations of homocysteine from 5.51 microM with no vitamin supplement to 4.54 and 4.77 +/- 0.37 microM, respectively. Results of the present experiment suggest that the effects of the combined

  7. Oxidation of methionine 216 in sheep and elk prion protein is highly dependent upon the amino acid at position 218 but is not important for prion propagation.

    PubMed

    Silva, Christopher J; Dynin, Irina; Erickson, Melissa L; Requena, Jesús R; Balachandran, Aru; Hui, Colleen; Onisko, Bruce C; Carter, John Mark

    2013-03-26

    We employed a sensitive mass spectrometry-based method to deconstruct, confirm, and quantitate the prions present in elk naturally infected with chronic wasting disease and sheep naturally infected with scrapie. We used this approach to study the oxidation of a methionine at position 216 (Met216), because this oxidation (MetSO216) has been implicated in prion formation. Three polymorphisms (Ile218, Val218, and Thr218) of sheep recombinant prion protein were prepared. Our analysis showed the novel result that the proportion of MetSO216 was highly dependent upon the amino acid residue at position 218 (I > V > T), indicating that Ile218 in sheep and elk prion protein (PrP) renders the Met216 intrinsically more susceptible to oxidation than the Val218 or Thr218 analogue. We were able to quantitate the prions in the attomole range. The presence of prions was verified by the detection of two confirmatory peptides: GENFTETDIK (sheep and elk) and ESQAYYQR (sheep) or ESEAYYQR (elk). This approach required much smaller amounts of tissue (600 μg) than traditional methods of detection (enzyme-linked immunosorbent assay, Western blot, and immunohistochemical analysis) (60 mg). In sheep and elk, a normal cellular prion protein containing MetSO216 is not actively recruited and converted to prions, although we observed that this Met216 is intrinsically more susceptible to oxidation.

  8. Geochemical behaviour of palladium in soils and Pd/PdO model substances in the presence of the organic complexing agents L-methionine and citric acid.

    PubMed

    Zereini, Fathi; Wiseman, Clare L S; Vang, My; Albers, Peter; Schneider, Wolfgang; Schindl, Roland; Leopold, Kerstin

    2016-01-01

    Risk assessments of platinum group metal (PGE) emissions, notably those of platinum (Pt), palladium (Pd) and rhodium (Rh), have been mostly based on data regarding the metallic forms used in vehicular exhaust converters, known to be virtually biologically inert and immobile. To adequately assess the potential impacts of PGE, however, data on the chemical behaviour of these metals under ambient conditions post-emission is needed. Complexing agents with a high affinity for metals in the environment are hypothesized to contribute to an increased bioaccessibility of PGE. The purpose of this study is to examine the modulating effects of the organic complexing agents, L-methionine and citric acid, on the geochemical behavior of Pd in soils and model substances (Pd black and PdO). Batch experimental tests were conducted with soils and model substances to examine the impacts of the concentration of complexing agents, pH and length of extraction period on Pd solubility and its chemical transformation. Particle surface chemistry was examined using X-ray photoelectron spectroscopy (XPS) on samples treated with solutions under various conditions, including low and high O2 levels. Pd was observed to be more soluble in the presence of organic complexing agents, compared to Pt and Rh. Pd in soils was more readily solubilized with organic complexing agents compared to the model substances. After 7 days of extraction, L-methionine (0.1 M) treated soil and Pd black samples, for instance, had mean soluble Pd fractions of 12.4 ± 5.9% and 0.554 ± 0.024%, respectively. Surface chemistry analyses (XPS) confirmed the oxidation of metallic Pd surfaces when treated with organic complexing agents. The type of organic complexing agent used for experimental purposes was observed to be the most important factor influencing solubility, followed by solution pH and time of extraction. The results demonstrate that metallic Pd can be transformed into more bioaccessible species in the presence of

  9. Overexpression of wild-type aspartokinase increases L-lysine production in the thermotolerant methylotrophic bacterium Bacillus methanolicus.

    PubMed

    Jakobsen, Oyvind M; Brautaset, Trygve; Degnes, Kristin F; Heggeset, Tonje M B; Balzer, Simone; Flickinger, Michael C; Valla, Svein; Ellingsen, Trond E

    2009-02-01

    Aspartokinase (AK) controls the carbon flow into the aspartate pathway for the biosynthesis of the amino acids l-methionine, l-threonine, l-isoleucine, and l-lysine. We report here the cloning of four genes (asd, encoding aspartate semialdehyde dehydrogenase; dapA, encoding dihydrodipicolinate synthase; dapG, encoding AKI; and yclM, encoding AKIII) of the aspartate pathway in Bacillus methanolicus MGA3. Together with the known AKII gene lysC, dapG and yclM form a set of three AK genes in this organism. Overexpression of dapG, lysC, and yclM increased l-lysine production in wild-type B. methanolicus strain MGA3 2-, 10-, and 60-fold (corresponding to 11 g/liter), respectively, without negatively affecting the specific growth rate. The production levels of l-methionine (less than 0.5 g/liter) and l-threonine (less than 0.1 g/liter) were low in all recombinant strains. The AK proteins were purified, and biochemical analyses demonstrated that they have similar V(max) values (between 47 and 58 micromol/min/mg protein) and K(m) values for l-aspartate (between 1.9 and 5.0 mM). AKI and AKII were allosterically inhibited by meso-diaminopimelate (50% inhibitory concentration [IC(50)], 0.1 mM) and by l-lysine (IC(50), 0.3 mM), respectively. AKIII was inhibited by l-threonine (IC(50), 4 mM) and by l-lysine (IC(50), 5 mM), and this enzyme was synergistically inhibited in the presence of both of these amino acids at low concentrations. The correlation between the impact on l-lysine production in vivo and the biochemical properties in vitro of the individual AK proteins is discussed. This is the first example of improving l-lysine production by metabolic engineering of B. methanolicus and also the first documentation of considerably increasing l-lysine production by overexpression of a wild-type AK.

  10. N-Terminal Methionine Processing.

    PubMed

    Wingfield, Paul T

    2017-04-03

    Protein synthesis is initiated by methionine in eukaryotes and by formylmethionine in prokaryotes. N-terminal methionine can be co-translationally cleaved by the enzyme methionine aminopeptidase (MAP). When recombinant proteins are expressed in bacterial and mammalian expression systems, there is a simple universal rule that predicts whether the initiating methionine will be processed by MAP based on the size of the residue adjacent (penultimate) to the N-methionine. In general, if the side chains of the penultimate residues have a radius of gyration of 1.29 Å or less, methionine is cleaved. © 2017 by John Wiley & Sons, Inc.

  11. Methionine-to-Cysteine Recycling in Klebsiella aerogenes

    PubMed Central

    Seiflein, Thomas A.; Lawrence, Jeffrey G.

    2001-01-01

    In the enteric bacteria Escherichia coli and Salmonella enterica, sulfate is reduced to sulfide and assimilated into the amino acid cysteine; in turn, cysteine provides the sulfur atom for other sulfur-bearing molecules in the cell, including methionine. These organisms cannot use methionine as a sole source of sulfur. Here we report that this constraint is not shared by many other enteric bacteria, which can use either cysteine or methionine as the sole source of sulfur. The enteric bacterium Klebsiella aerogenes appears to use at least two pathways to allow the reduced sulfur of methionine to be recycled into cysteine. In addition, the ability to recycle methionine on solid media, where cys mutants cannot use methionine as a sulfur source, appears to be different from that in liquid media, where they can. One pathway likely uses a cystathionine intermediate to convert homocysteine to cysteine and is induced under conditions of sulfur starvation, which is likely sensed by low levels of the sulfate reduction intermediate adenosine-5′-phosphosulfate. The CysB regulatory proteins appear to control activation of this pathway. A second pathway may use a methanesulfonate intermediate to convert methionine-derived methanethiol to sulfite. While the transsulfurylation pathway may be directed to recovery of methionine, the methanethiol pathway likely represents a general salvage mechanism for recovery of alkane sulfide and alkane sulfonates. Therefore, the relatively distinct biosyntheses of cysteine and methionine in E. coli and Salmonella appear to be more intertwined in Klebsiella. PMID:11114934

  12. Post-translational enzyme modification by the phosphopantetheinyl transferase is required for lysine and penicillin biosynthesis but not for roquefortine or fatty acid formation in Penicillium chrysogenum.

    PubMed

    García-Estrada, Carlos; Ullán, Ricardo V; Velasco-Conde, Tania; Godio, Ramiro P; Teijeira, Fernando; Vaca, Inmaculada; Feltrer, Raúl; Kosalková, Katarina; Mauriz, Elba; Martín, Juan F

    2008-10-15

    NRPSs (non-ribosomal peptide synthetases) and PKSs (polyketide synthases) require post-translational phosphopantetheinylation to become active. This reaction is catalysed by a PPTase (4'-phosphopantetheinyl transferase). The ppt gene of Penicillium chrysogenum, encoding a protein that shares 50% similarity with the stand-alone large PPTases, has been cloned. This gene is present as a single copy in the genome of the wild-type and high-penicillin-producing strains (containing multiple copies of the penicillin gene cluster). Amplification of the ppt gene produced increases in isopenicillin N and benzylpenicillin biosynthesis. A PPTase-defective mutant (Wis54-PPT(-)) was obtained. It required lysine and lacked pigment and penicillin production, but it still synthesized normal levels of roquefortine. The biosynthesis of roquefortine does not appear to involve PPTase-mediated modification of the synthesizing enzymes. The PPT(-) mutant did not require fatty acids, which indicates that activation of the fatty acid synthase is performed by a different PPTase. Complementation of Wis54-PPT(-) with the ppt gene restored lysine biosynthesis, pigmentation and penicillin production, which demonstrates the wide range of processes controlled by this gene.

  13. Physiological and biochemical aspects of methionine isomers and a methionine analogue in broilers.

    PubMed

    Zhang, Shuai; Saremi, Behnam; Gilbert, Elizabeth R; Wong, Eric A

    2017-02-01

    Methionine is the first limiting amino acid in all poultry corn-soybean based diets. The objective of this study was to determine the effect of supplementation of L-methionine (L-Met), DL-methionine (DL-Met), and the methionine analogue, DL-2-hydroxy-4-(methylthio) butanoic acid (DL-HMTBA), on biochemical and physiological parameters of broiler chickens. Male Cobb-500 broilers were fed from day of hatch (d 0) to d 35 posthatch using a basal diet deficient in methionine plus cysteine (Met + Cys) (control), or the basal diet supplemented with 0.22% DL-Met, 0.22% L-Met, or 0.31% DL-HMTBA to meet the Met + Cys requirements. Tissue (liver, duodenum, jejunum, and ileum) and blood samples were collected at various ages, from d 0 to d 35. Performance of the birds, blood parameters (e.g., acute phase proteins, white blood cell counts), mRNA expression of intestinal nutrient transporters and DNA methylation properties of liver tissues were examined. Both body weight and feed efficiency were improved in methionine supplemented groups compared to the control group. No significant differences were observed among DL-Met, L-Met, and DL-HMTBA for growth performance parameters. L-Met and DL-Met supplementation decreased the acute phase protein, serum amyloid A, while DL-HMTBA had no effect. Methionine supplementation had no effect on white blood cell differentiation count, hepatic total DNA methylation, or DNA methyltransferase activity. L-Met and DL-Met, but not DL-HMTBA, supplementation, resulted in enhanced expression of the ATB(0,+) and B(0)AT transporters in various small intestinal segments. All methionine sources increased expression of MCT1 in the jejunum. In conclusion, methionine supplementation improved growth performance of male broilers. Methionine supplementation was also associated with changes in intestinal nutrient transporter gene expression in certain segments and ages, suggesting that intestinal amino acid absorptive function can be regulated by the source

  14. NUTRITIONAL FACTORS STIMULATING THE FORMATION OF LYSINE DECARBOXYLASE IN ESCHERICHIA COLI

    PubMed Central

    Maretzki, Andrew; Mallette, M. F.

    1962-01-01

    Maretzki, Andrew (Pennsylvania State University, University Park) and M. F. Mallette. Nutritional factors stimulating the formation of lysine decarboxylase in Escherichia coli. J. Bacteriol. 83:720–726. 1962 — Inclusion of complex nitrogen sources in the induction medium was shown to be necessary for the synthesis of appreciable amounts of l-lysine decarboxylase by Escherichia coli B. Hy-case, a commercial acid hydrolyzate of casein, was especially effective in enzyme production, which was assayed manometrically after lysis of the bacteria from without by bacteriophage. Partial fractionation of the Hy-case, identification of the free amino acids, and addition of these amino acids to test media revealed stimulatory effects by methionine, threonine, proline, leucine, and tyrosine. A full complement of amino acids did not match the enzyme levels reached in the presence of Hy-case. Certain peptide fractions obtained from this mixture supplemented the effects of the amino acids in such a way as to suggest direct incorporation of peptide rather than transport or protective roles. Added purines, pyrimidines, iron, and water-soluble vitamins were without effect. Neither carbohydrates nor phosphorylated materials could be detected in the stimulatory fractions. PMID:14469751

  15. Controlled expression and structural organization of a Lactococcus lactis bacteriophage lysin encoded by two overlapping genes.

    PubMed Central

    Shearman, C A; Jury, K L; Gasson, M J

    1994-01-01

    The phi vML3 bacteriophage lysin is specific for lactococci and could be used to promote enzyme release during cheese manufacture. The level of lysin expression from the cloned gene using its own upstream sequences is very low. Expression in Escherichia coli by using a synthetic hybrid lysin gene and a series of BAL 31 deletions of the original cloned DNA fragment suggested that the start of the gene had previously been incorrectly assigned. Reevaluation of homology between the lysin and Bacillus subtilis PZA protein 15 led to the identification of a new potential ribosome binding site (RBS). A 0.72-kb PCR-generated fragment including this RBS and the complete lysin gene was expressed and inducibly controlled. The translational start of the lysin gene was identified as an isoleucine codon, and this may lead to a low translation rate. During the analysis of the BAL 31 deletion fragments, two proteins of 20 and 8 kDa were shown to be expressed from the originally defined lysin gene. The DNA sequence has a second open reading frame with a good RBS and two potential start methionines. The smaller lysin protein was isolated, and the N terminus was sequenced, confirming that one methionine codon acted as the start of a second gene. The larger lysin protein has homology with lysozymes. The smaller lysin protein has some features resembling those of a holin. The possible roles of these two proteins in lysis of lactococci are discussed. Images PMID:7944354

  16. Dissimilation of Methionine by Achromobacter starkeyi1

    PubMed Central

    Ruiz-Herrera, José; Starkey, Robert L.

    1970-01-01

    Methionine was decomposed by some bacteria which were isolated from soil. The sulfur of the methionine was liberated as methanethiol, and part of this became oxidized to dimethyl disulfide. Detailed studies with one of these cultures, Achromobacter starkeyi, indicated that the first step in methionine decomposition was its oxidadative deamination to α-keto-γ-methyl mercaptobutyrate by a constitutive amino acid oxidase. The following steps were carried out by inducible enzymes, the synthesis of which was inhibited by chloramphenicol. α-Keto-γ-methyl mercaptobutyrate was split producing methanethiol and α-keto butyrate which was oxidized to propionate. The metabolism of propionate was similar to that described for animal tissues; the propionate was carboxylated to succinate via methyl malonyl coenzyme A, and the succinate was metabolized through the Krebs cycle. PMID:16559105

  17. Synthesis and Anchoring of Antineoplastic Ferrocene and Phthalocyanine Derivatives on Water-Soluble Polymeric Drug Carriers Derived from Lysine and Aspartic Acid

    PubMed Central

    Maree, M. David; Neuse, Eberhard W.; Erasmus, Elizabeth; Swarts, Jannie C.

    2008-01-01

    The general synthetic strategy towards water-soluble biodegradable drug carriers and the properties that they must have are discussed. The syntheses of water-soluble biodegradable copolymers of lysine and aspartic acid as potential drug-delivering devices, having amine-functionalised side chains are then described. Covalent anchoring of carboxylic acid derivatives of the antineoplastic ferrocene and photodynamically active phthalocyanine moieties to the amine-containing drug carrier copolymers under mild coupling conditions has been achieved utilising the coupling reagent O-benzotriazolyl-N,N,N′,N′-tetramethyluronium hexafluorophosphate to promote formation of the biodegradable amide bond. Even though the parent antineoplastic ferrocene and phthalocyanine derivatives are themselves insoluble in water at pH < 7, the new carrier-drug conjugates that were obtained are well water-soluble. PMID:18288243

  18. Methionine and methionine sulfoxide treatment induces M1/classical macrophage polarization and modulates oxidative stress and purinergic signaling parameters.

    PubMed

    Dos Santos, Lien M; da Silva, Tatiane M; Azambuja, Juliana H; Ramos, Priscila T; Oliveira, Pathise S; da Silveira, Elita F; Pedra, Nathalia S; Galdino, Kennia; do Couto, Carlus A T; Soares, Mayara S P; Tavares, Rejane G; Spanevello, Roselia M; Stefanello, Francieli M; Braganhol, Elizandra

    2017-01-01

    Methionine is an essential amino acid involved in critical metabolic process, and regulation of methionine flux through metabolism is important to supply this amino acid for cell needs. Elevation in plasma methionine commonly occurs due to mutations in methionine-metabolizing enzymes, such as methionine adenosyltransferase. Hypermethioninemic patients exhibit clinical manifestations, including neuronal and liver disorders involving inflammation and tissue injury, which pathophysiology is not completely established. Here, we hypothesize that alterations in macrophage inflammatory response may contribute to deleterious effects of hypermethioninemia. To this end, macrophage primary cultures were exposed to methionine (1 mM) and/or its metabolite methionine sulfoxide (0.5 mM), and M1/proinflammatory or M2/anti-inflammatory macrophage polarization was evaluated. In addition, inflammation-related pathways including oxidative stress parameters, as superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPx) activities; reactive oxygen species (ROS) production, and purinergic signaling, as ATP/ADP/AMPase activities, were investigated. Methionine and/or methionine sulfoxide induced M1/classical macrophage activation, which is related to proinflammatory responses characterized by increased iNOS activity and TNF-α release. Further experiments showed that treatments promoted alterations on redox state of macrophages by differentially modulated SOD and CAT activities and ROS levels. Finally, methionine and/or methionine sulfoxide treatment also altered the extracellular nucleotide metabolism, promoting an increase of ATPase/ADPase activities in macrophages. In conclusion, these findings contribute to better understand the participation of proinflammatory responses in cell injury observed in hypermethioninemic patients.

  19. A study of quenching effect of sulfur-containing amino acids L-cysteine and L-methionine on peroxyoxalate chemiluminescence of 7-amino-4-trifluoromethylcumarin

    NASA Astrophysics Data System (ADS)

    Shamsipur, Mojtaba; Chaichi, Mohammad Javad

    2005-04-01

    The quenching effect of L-cysteine and L-methionine on strong chemiluminescence of bis-(2,4,6-trichlorophenyl)oxalate-H 2O 2 system in the presence of 7-amino-4-trifluoromethylcumarin was studied. The chemiluminescence parameters were evaluated from computer fitting of the resulting intensity-time plots. Both systems resulted in Stern-Volmer plots in the quencher concentration range of 6.5 × 10 -5 to 5.2 × 10 -4 M with kQ values of 1.15 × 10 4 and 1.28 × 10 4 M -1 for L-cysteine and L-methionine, respectively.

  20. Lysine Fermentation: History and Genome Breeding.

    PubMed

    Ikeda, Masato

    2016-11-11

    Lysine fermentation by Corynebacterium glutamicum was developed in 1958 by Kyowa Hakko Kogyo Co. Ltd. (current Kyowa Hakko Bio Co. Ltd.) and is the second oldest amino acid fermentation process after glutamate fermentation. The fundamental mechanism of lysine production, discovered in the early stages of the process's history, gave birth to the concept known as "metabolic regulatory fermentation," which is now widely applied to metabolite production. After the development of rational metabolic engineering, research on lysine production first highlighted the need for engineering of the central metabolism from the viewpoints of precursor supply and NADPH regeneration. Furthermore, the existence of active export systems for amino acids was first demonstrated for lysine in C. glutamicum, and this discovery has resulted in the current recognition of such exporters as an important consideration in metabolite production. Lysine fermentation is also notable as the first process to which genomics was successfully applied to improve amino acid production. The first global "genome breeding" strategy was developed using a lysine producer as a model; this has since led to new lysine producers that are more efficient than classical industrial producers. These advances in strain development technology, combined with recent systems-level approaches, have almost achieved the optimization of entire cellular systems as cell factories for lysine production. In parallel, the continuous improvement of the process has resulted not only in fermentation processes with reduced load on downstream processing but also in commercialization of various product forms according to their intended uses. Nowadays lysine fermentation underpins a giant lysine demand of more than 2 million metric tons per year.

  1. Oxidation of methionine in PrP is dependent upon the oxidant and the amino acid two positions removed(Abstract)

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Background/Introduction. Methionine oxidation has been shown both to be associated with prion formation and implicated in the inhibition of amyloid formation in model systems. This work is based on model systems where hydrogen peroxide was used as an oxidant. Materials and Methods. We developed a se...

  2. A phase transition from monoclinic C2 with Z' = 1 to triclinic P1 with Z' = 4 for the quasiracemate L-2-aminobutyric acid-D-methionine (1/1).

    PubMed

    Görbitz, Carl Henrik; Wragg, David S; Bakke, Ingrid Marie Bergh; Fleischer, Christian; Grønnevik, Gaute; Mykland, Maria; Park, Yoomin; Trovik, Kristian Wiedicke; Serigstad, Halvard; Sundsli, Bård Edgar Vestheim

    2016-07-01

    Racemates of hydrophobic amino acids with linear side chains are known to undergo a unique series of solid-state phase transitions that involve sliding of molecular bilayers upon heating or cooling. Recently, this behaviour was shown to extend also to quasiracemates of two different amino acids with opposite handedness [Görbitz & Karen (2015). J. Phys. Chem. B, 119, 4975-4984]. Previous investigations are here extended to an L-2-aminobutyric acid-D-methionine (1/1) co-crystal, C4H9NO2·C5H11NO2S. The significant difference in size between the -CH2CH3 and -CH2CH2SCH3 side chains leads to extensive disorder at room temperature, which is essentially resolved after a phase transition at 229 K to an unprecedented triclinic form where all four D-methionine molecules in the asymmetric unit have different side-chain conformations and all three side-chain rotamers are used for the four partner L-2-aminobutyric acid molecules.

  3. Lysine catabolism in Rhizoctonia leguminicola and related fungi.

    PubMed Central

    Guengerich, F P; Broquist, H P

    1976-01-01

    The catabolism of lysine was studied in several yeasts and fungi. Results with cell-free extracts of Rhizoctonia leguminicola support a proposed pathway involving (D- and L-) EPSILON-N-acetyllysine, alpha-keto-epsilon-acetamidohexanoic acid, delta-acetamidovaleric acid, and delta-aminovaleric acid in the conversion of L-lysine to shortchain organic acids. Label from radioactive L-lysine was found to accumulate in D- and L-epsilon-N-acetyllysine, delta-acetamidovaleric acid, delta-aminovaleric acid, and glutaric acid in cultures of R. leguminicola, Neurospora crassa, Saccharomyces cerevisiae, and Hansenula saturnus, suggesting that the proposed omega-acetyl pathway of lysine catabolism is generalized among yeasts and fungi. In N. crassa, as is the case in R. leguminicola, the major precursor of L-pipecolic acid was the L-isomer of lysine; 15N experiments were consistent with delta1-piperideine-2-carboxylic acid as an intermediate in the transformation. PMID:131119

  4. Lysine catabolism in Rhizoctonia leguminicola and related fungi.

    PubMed

    Guengerich, F P; Broquist, H P

    1976-04-01

    The catabolism of lysine was studied in several yeasts and fungi. Results with cell-free extracts of Rhizoctonia leguminicola support a proposed pathway involving (D- and L-) EPSILON-N-acetyllysine, alpha-keto-epsilon-acetamidohexanoic acid, delta-acetamidovaleric acid, and delta-aminovaleric acid in the conversion of L-lysine to shortchain organic acids. Label from radioactive L-lysine was found to accumulate in D- and L-epsilon-N-acetyllysine, delta-acetamidovaleric acid, delta-aminovaleric acid, and glutaric acid in cultures of R. leguminicola, Neurospora crassa, Saccharomyces cerevisiae, and Hansenula saturnus, suggesting that the proposed omega-acetyl pathway of lysine catabolism is generalized among yeasts and fungi. In N. crassa, as is the case in R. leguminicola, the major precursor of L-pipecolic acid was the L-isomer of lysine; 15N experiments were consistent with delta1-piperideine-2-carboxylic acid as an intermediate in the transformation.

  5. Efficiency of lysine or threonine retention in growing rats fed diets limiting in either lysine or threonine.

    PubMed

    Gahl, M J; Finke, M D; Crenshaw, T D; Benevenga, N J

    1996-12-01

    Over a 21-d experiment, the efficiency of lysine and threonine retention was determined in 80 male Sprague-Dawley rats (65.9 +/- 0.3 g, means +/- SE) fed purified diets containing an amino acid mix limiting in either lysine or threonine. With additional increments of the first limiting amino acid, lysine concentration in total body protein (g/16 g N) increased (P < 0.01) in rats fed lysine-limiting diets but, when fed threonine-limiting diets, lysine concentration in body protein first increased and then decreased (P < 0.01). As increments of the first limiting amino acid were added, the threonine concentration in total body protein increased then decreased when both lysine- (P < 0.01) and threonine- (P < 0.06) limiting diets were fed. Lysine and threonine retention were calculated based on comparative slaughter. Sixteen rats were killed on d 0 to estimate the grams of amino acid in the body. Retention responses were analyzed using a logistic equation in which lysine or threonine intake was used to predict retention. The maximum marginal efficiency (dr/dI, retention/intake) was observed at <40% of maximum retention. For lysine retention, it was 81% when lysine was limiting and 70% when threonine was limiting. For threonine retention, it was 58% when threonine was limiting and 49% when lysine was limiting. The maximum cumulative efficiency (retention adjusted for maintenance relative to cumulative intake) for lysine retention was 62% when lysine was limiting or 58% when threonine was limiting. For threonine retention, it was 51% when threonine was limiting and 35% when lysine was limiting. Thus, amino acid concentration in body protein is not constant, and amino acids are used with higher efficiency when first limiting.

  6. Adaptation of human immunodeficiency virus type 1 to cells expressing a binding-deficient CD4 mutant (lysine 46 to aspartic acid).

    PubMed Central

    Choe, H R; Sodroski, J

    1995-01-01

    Human immunodeficiency virus (HIV-1) was adapted to replicate efficiently in cells expressing an altered form of the CD4 viral receptor. The mutant CD4 (46 K/D) contained a single amino acid change (lysine 46 to aspartic acid) in the CDR2 loop of domain 1, which results in a 15-fold reduction in affinity for the viral gp120 glycoprotein. The ability of the adapted virus to replicate in CD4 46 K/D-expressing cells was independently enhanced by single amino acid changes in the V2 variable loop, the V3 variable loop, and the fourth conserved (C4) region of the gp120 glycoprotein. Combinations of these amino acids in the same envelope glycoprotein resulted in additive enhancement of virus replication in cells expressing the CD4 46 K/D molecule. In cells expressing the wild-type CD4 glycoproteins, the same V2 and V3 residue changes also increased the efficiency of replication of a virus exhibiting decreased receptor-binding ability due to an amino acid change (aspartic acid 368 to glutamic acid) in the gp120 glycoprotein. In neither instance did the adaptive changes restore the binding ability of the monomeric gp120 glycoprotein or the oligomeric envelope glycoprotein complex for the mutant or wild-type CD4 glycoproteins, respectively. Thus, particular conformations of the gp120 V2 and V3 variable loops and of the C4 region allow postreceptor binding events in the membrane fusion process to occur in the context of less than optimal receptor binding. These results suggest that the fusion-related functions of the V2, V3, and C4 regions of gp120 are modulated by CD4 binding. PMID:7707502

  7. Estimation of optimum amino acid supplements to triticale.

    PubMed

    Heger, J

    1990-04-01

    Based on the nitrogen balance (NB) data and the efficiency of amino acid utilization, optimum supplements of limiting amino acids (AA) to triticale were calculated and their effect on true N digestibility (TD), biological value of protein (BV) and net protein utilization (NPU) of a triticale-based diet was evaluated. To determine if AA balance must be considered in the calculation of the optimum supplements, the effect of a 20% or 40% excess of essential AA in lysine-, methionine-, threonine- or tryptophan-deficient diets was also studied. The AA excess had no significant effect on NB in diets deficient in lysine, methionine or threonine. However, NB in rats fed on the tryptophan-deficient diet increased as the AA excess increased. BV of the diet containing the optimized supplements of lysine, threonine, methionine and valine was comparable to that of lactalbumin or to the diet supplemented with all essential AA. The deletion of valine from the optimized supplement caused an insignificant decrease in BV. Due to the lower TD, NPU of diets containing the optimized AA supplements was lower than that of the diet containing all essential AA or of the lactalbumin-based diet.

  8. Economic process to co-produce poly(ε-l-lysine) and poly(l-diaminopropionic acid) by a pH and dissolved oxygen control strategy.

    PubMed

    Xu, Zhaoxian; Feng, Xiaohai; Sun, Zhuzhen; Cao, Changhong; Li, Sha; Xu, Zheng; Xu, Zongqi; Bo, Fangfang; Xu, Hong

    2015-01-01

    This study tended to apply biorefinery of indigenous microbes to the fermentation of target-product generation through a novel control strategy. A novel strategy for co-producing two valuable homopoly(amino acid)s, poly(ε-l-lysine) (ε-PL) and poly(l-diaminopropionic acid) (PDAP), was developed by controlling pH and dissolved oxygen concentrations in Streptomyces albulus PD-1 fermentation. The production of ε-PL and PDAP got 29.4 and 9.6gL(-1), respectively, via fed-batch cultivation in a 5L bioreactor. What is more, the highest production yield (21.8%) of similar production systems was achieved by using this novel strategy. To consider the economic-feasibility, large-scale production in a 1t fermentor was also implemented, which would increase the gross profit of 54,243.5USD from one fed-batch bioprocess. This type of fermentation, which produces multiple commercial products from a unified process is attractive, because it will improve the utilization rate of raw materials, enhance production value and enrich product variety.

  9. A pH-responsive drug nanovehicle constructed by reversible attachment of cholesterol to PEGylated poly(l-lysine) via catechol-boronic acid ester formation.

    PubMed

    Yang, Bin; Lv, Yin; Zhu, Jing-Yi; Han, Yun-Tao; Jia, Hui-Zhen; Chen, Wei-Hai; Feng, Jun; Zhang, Xian-Zheng; Zhuo, Ren-Xi

    2014-08-01

    The present work reports the construction of a drug delivery nanovehicle via a pH-sensitive assembly strategy for improved cellular internalization and intracellular drug liberation. Through spontaneous formation of boronate linkage in physiological conditions, phenylboronic acid-modified cholesterol was able to attach onto catechol-pending methoxypoly(ethylene glycol)-block-poly(l-lysine). This comb-type polymer can self-organize into a micellar nanoconstruction that is able to effectively encapsulate poorly water-soluble agents. The blank micelles exhibited negligible in vitro cytotoxicity, yet doxorubicin (DOX)-loaded micelles could effectively induce cell death at a level comparable to free DOX. Owing to the acid-labile feature of the boronate linkage, a reduction in environmental pH from pH 7.4 to 5.0 could trigger the dissociation of the nanoconstruction, which in turn could accelerate the liberation of entrapped drugs. Importantly, the blockage of endosomal acidification in HeLa cells by NH4Cl treatment significantly decreased the nuclear uptake efficiency and cell-killing effect mediated by the DOX-loaded nanoassembly, suggesting that acid-triggered destruction of the nanoconstruction is of significant importance in enhanced drug efficacy. Moreover, confocal fluorescence microscopy and flow cytometry assay revealed the effective internalization of the nanoassemblies, and their cellular uptake exhibited a cholesterol dose-dependent profile, indicating the contribution of introduced cholesterol functionality to the transmembrane process of the nanoassembly.

  10. [Changes in the lysine of spiruline algae samples after various heat treatments].

    PubMed

    Adrian, J

    1975-01-01

    The spirulina algae are microorganisms which are cultivated on Mexican lakes for feeding use. After drying, they contain about 52 per cent of proteins, with 4 per cent of lysine and 1, 7 per cent of methionine. In the studied samples, pH is 6, 2; they are partially autolysed and contain 4 to 18 per cent of free lysine and methionine and 6, 5 per cent of soluble carbohydrates. During heating treatments, the spiurlina lysine reacts as the same as oilcak meal lysine; it resists rather well to autoclaving but less to roasting. The yeast lysine is more stable than the spirulina lysine. The thermic stability of spirulina lysine is caused first by the small amount of free reducing carbohydrates, and for a minor part by the natural acicity of these products. The lysine destruction is proportional to the autolysis stage of the samples, that is to say the presence of free aminoacids. All the behavior differences between the various spirulina samples disappear when are heated with xylose, which induces a strong Maillard reaction.

  11. In Vitro Synthesis of a Precursor to the Methionine-rich Polypeptide of the Zein Fraction of Corn 1

    PubMed Central

    Melcher, Ulrich

    1979-01-01

    The messenger ribonucleic acid fraction isolated from a protein bodyenriched fraction of developing corn (Zea mays L.) endosperm stimulated the incorporation of radioactive amino acids into at least five polypeptides when added to a wheat germ extract capable of protein synthesis. Of these, the two major polypeptides formed with messenger from freshly frozen corn were identified as precursors to zein A and B, the two major polypeptides of the prolamine fraction of corn meal (21,600 and 19,600 molecular weight). The identification was based on the relative incorporations of radioactive leucine, lysine, and methionine, and the susceptibility of the zein A precursor, but not the zein B precursor to cleavage with cyanogen bromide. Using extracts from stored frozen corn, a third polypeptide of 14,500 molecular weight was identified as a major in vitro product. It was preferentially labeled with methionine and slightly larger than a similar peptide in the zein fraction of corn meal. Two other polypeptides of still lower molecular weight could be detected above the background of probably incomplete polypeptides. Images PMID:16660727

  12. Bioavailability of different dietary supplemental methionine sources in animals.

    PubMed

    Zhang, Shuai; Wong, Eric A; Gilbert, Elizabeth R

    2015-06-01

    Dietary methionine is indispensable for animal maintenance, growth and development. L-methionine (L-Met), and its synthetic forms DL-methionine (DL-Met) and 2-hydroxy-4 (methylthio) butanoic acid (HMTBA) are common supplemental methionine sources in animal diets. There are different characteristics for cellular absorption, transport, metabolism and bio-efficiency between these three dietary methionine sources. Moreover, there are differences in their utilization among various species such as chickens, pigs and ruminants. As a methionine precursor, HMTBA is efficacious in the promotion of growth in animals. It is absorbed mainly by monocarboxylate transporter 1 (MCT1), coupled with the activity of the Na(+)/H(+) exchanger (NHE3), while DL-Met uptake occurs via multiple carrier-mediated systems. Liver, kidney and small intestine can metabolize D-Met and HMTBA to L-Met through oxidation and transamination. In ruminants, the non-hepatic tissues act as major sites of HMTBA conversion, which are different from that in chickens and pigs. HMTBA also has additional benefits in anti-oxidation. Understanding the characteristics of uptake and metabolism of different methionine sources will greatly benefit the industry and bioscience research.

  13. Protein-borne methionine residues as structural antioxidants in mitochondria.

    PubMed

    Schindeldecker, Mario; Moosmann, Bernd

    2015-07-01

    Methionine is an oxidant-labile amino acid whose major oxidation products, methionine sulfoxides, can be readily repaired by various NADPH-dependent methionine sulfoxide reductases. Formally, the methionine oxidation-reduction circuit could act as a cellular antioxidant system, by providing a safe sink for oxidants that might cause much more damage if reacting otherwise. This concept is supported by focal experimental evidence; however, the global importance, scope and biochemical role of protein-borne methionine as an inbuilt macromolecular antioxidant have remained incompletely defined. In analyzing proteomic methionine usage on different levels of comparison, we find that protein methionine (i) is primarily an antioxidant of mitochondria, especially of the inner mitochondrial membrane, (ii) responds strongly to respiratory demands on an evolutionary timescale, (iii) acts locally, by selectively protecting its carrier protein, and (iv) might be utilized as a molecular predictor of aerobic metabolic rate in animals, to complement traditional markers like the presence of a respiratory pigment. Our data support the idea that proteins in need of a long lifespan or acting in dangerous environments may acquire massive structural alterations aimed at increasing their resistance to oxidation. Counterintuitively though, they sometimes do so by accumulating particularly labile rather than particularly stable building blocks, illustrating that the technical concept of cathodic protection is also employed by the animate nature.

  14. Economical production of poly(ε-l-lysine) and poly(l-diaminopropionic acid) using cane molasses and hydrolysate of streptomyces cells by Streptomyces albulus PD-1.

    PubMed

    Xia, Jun; Xu, Zhaoxian; Xu, Hong; Liang, Jinfeng; Li, Sha; Feng, Xiaohai

    2014-07-01

    Poly(ε-L-lysine) (ε-PL) and poly(L-diaminopropionic acid) (PDAP) co-production by Streptomyces albulus PD-1 from cane molasses and hydrolysate of strepyomyces cells (HSC) was investigated for the first time in this study. The optimal initial total sugar concentration of the cane molasses pretreated with sulfuric acid was determined to be 20 g L(-1), and HSC could substitute for yeast extract for ε-PL and PDAP co-production. When fed-batch fermentation was performed in 1t fermentor with pretreated cane molasses and HSC, 20.6 ± 0.5 g L(-1) of ε-PL and 5.2 ± 0.6 g L(-1) of PDAP were obtained. The amount of strepyomyces cells obtained in one fed-batch fermentation is sufficient to prepare the HSC to satisfy the demand of subsequent fermentations, thus the self-cycling of organic nitrogen source becomes available. These results suggest that the low-cost cane molasses and HSC can be used for the economical production of ε-PL and PDAP by S. albulus PD-1.

  15. Amino acid composition and chemical evaluation of protein quality of cereals as affected by insect infestation.

    PubMed

    Jood, S; Kapoor, A C; Singh, R

    1995-09-01

    A significant decrease in essential amino acids of wheat, maize and sorghum was observed due to grain infestation caused by mixed populations of Trogoderma granarium Everts and Rhizopertha dominica Fabricius (50:50). Non-essential amino acids were also adversely affected. Among the essential amino acids, maximum reduction was found in methionine, isoleucine and lysine in infested wheat, maize and sorghum grains, respectively. Lysine, with lowest chemical score in uninfested and infested grains of three cereals, is the first limiting amino acid. Insect infestation caused significant (p < 0.05) reduction in the chemical score of all the essential amino acids, yet did not change the position of first and second limiting amino acids in wheat and sorghum. However, in case of maize, isoleucine became the second limiting amino acid. Infested grains also showed substantial reduction in essential amino acid index, calculated biological value and requirement index.

  16. Supplementation of a maternal low-protein diet in rat pregnancy with folic acid ameliorates programming effects upon feeding behaviour in the absence of disturbances to the methionine-homocysteine cycle.

    PubMed

    Engeham, Sarah F; Haase, Andrea; Langley-Evans, Simon C

    2010-04-01

    Maternal protein restriction in rat pregnancy is associated with altered feeding behaviour in later life. When allowed to self-select their diet, rats subject to prenatal undernutrition show an increased preference for fatty foods. The main aim of the present study was to evaluate the contribution of folic acid in the maternal diet to programming of appetite, since disturbances of the folate and methionine-homocysteine cycles have been suggested to impact upon epigenetic regulation of gene expression and hence programme long-term physiology and metabolism. Pregnant rats were fed diets containing either 9 or 18 % casein by weight, with folate provided at either 1 or 5 mg/kg diet. Adult male animals exposed to low protein (LP) in fetal life exhibited increased preference for high-fat food. Providing the higher level of folate in the maternal diet prevented this effect of LP, but offspring of rats fed 18 % casein diet with additional folate behaved in a similar manner to LP-exposed animals. Among day 20 gestation fetuses, it was apparent that both protein restriction and maternal folate supplementation could have adverse effects upon placental growth. Examination of methionine-homocysteine and folate cycle intermediates, tissue glutathione concentrations and expression of mRNA for methionine synthase, DNA methyltransferase 1 and methyltetrahydrofolate reductase revealed no gross disturbances of folate and one-carbon metabolism in either maternal or fetal tissue. The present findings indicated that any role for DNA methylation in programming of physiology is not related to major perturbations of folate metabolism, and is likely to be gene-specific rather than genome-wide.

  17. Transsulfuration in an adult with hepatic methionine adenosyltransferase deficiency.

    PubMed Central

    Gahl, W A; Bernardini, I; Finkelstein, J D; Tangerman, A; Martin, J J; Blom, H J; Mullen, K D; Mudd, S H

    1988-01-01

    We investigated sulfur and methyl group metabolism in a 31-yr-old man with partial hepatic methionine adenosyltransferase (MAT) deficiency. The patient's cultured fibroblasts and erythrocytes had normal MAT activity. Hepatic S-adenosylmethionine (SAM) was slightly decreased. This clinically normal individual lives with a 20-30-fold elevation of plasma methionine (0.72 mM). He excretes in his urine methionine and L-methionine-d-sulfoxide (2.7 mmol/d), a mixed disulfide of methanethiol and a thiol bound to an unidentified group X, which we abbreviate CH3S-SX (2.1 mmol/d), and smaller quantities of 4-methylthio-2-oxobutyrate and 3-methylthiopropionate. His breath contains 17-fold normal concentrations of dimethylsulfide. He converts only 6-7 mmol/d of methionine sulfur to inorganic sulfate. This abnormally low rate is due not to a decreased flux through the primarily defective enzyme, MAT, since SAM is produced at an essentially normal rate of 18 mmol/d, but rather to a rate of homocysteine methylation which is abnormally high in the face of the very elevated methionine concentrations demonstrated in this patient. These findings support the view that SAM (which is marginally low in this patient) is an important regulator that helps to determine the partitioning of homocysteine between degradation via cystathionine and conservation by reformation of methionine. In addition, these studies demonstrate that the methionine transamination pathway operates in the presence of an elevated body load of that amino acid in human beings, but is not sufficient to maintain methionine levels in a normal range. PMID:3339126

  18. Nucleotide sequence of a Pseudomonas denitrificans 5.4-kilobase DNA fragment containing five cob genes and identification of structural genes encoding S-adenosyl-L-methionine: uroporphyrinogen III methyltransferase and cobyrinic acid a,c-diamide synthase.

    PubMed Central

    Crouzet, J; Cauchois, L; Blanche, F; Debussche, L; Thibaut, D; Rouyez, M C; Rigault, S; Mayaux, J F; Cameron, B

    1990-01-01

    A 5.4-kilobase DNA fragment carrying Pseudomonas denitrificans cob genes has been sequenced. The nucleotide sequence and genetic analysis revealed that this fragment carries five different cob genes (cobA to cobE). Four of these genes present the characteristics of translationally coupled genes. cobA has been identified as the structural gene of S-adenosyl-L-methionine:uroporphyrinogen III methyltransferase (SUMT) because the encoded protein has the same NH2 terminus and molecular weight as those determined for the purified SUMT. For the same reasons the cobB gene was shown to be the structural gene for cobyrinic acid a,c-diamide synthase. Genetic and biochemical data concerning cobC and cobD mutants suggest that the products of these genes are involved in the conversion of cobyric acid to cobinamide. PMID:2211520

  19. Excessive S-Adenosyl-L-Methionine-Dependent Methylation Increases Levels of Methanol, Formaldehyde and Formic Acid in Rat Brain Striatal Homogenates: Possible role in S-adenosyl-L-methionine-induced Parkinson’s disease-like disorders

    PubMed Central

    Lee, Eun-Sook; Chen, Hongtao; Hardman, Chadwick; Simm, Anthony; Charlton, Clivel

    2009-01-01

    Aims Excessive methylation may be a precipitating factor for Parkinson’s disease (PD) since S-adenosylmethionine (SAM), the endogenous methyl donor, induces PD-like changes when injected into the rat brain. The hydrolysis of the methyl ester bond of the methylated proteins produces methanol. Since methanol is oxidized into formaldehyde, and formaldehyde into formic acid in the body, we investigated the effects of SAM on the production of methanol, formaldehyde and formic acid in rat brain striatal homogenates and the toxicity of these products in PC12 cells. Main methods radio-enzymatic and colorimetric assays, cell viability, Western blot. Key findings SAM increased the formation of methanol, formaldehyde and formic acid in a concentration and time-dependent manner. Concentrations of [3H-methyl]-SAM at 0.17, 0.33, 0.67 and 1.34 nM produced 3.8, 8.0, 18.3 and 34.4 fmol/mg protein/h of [3H] methanol in rat striatal homogenates, respectively. SAM also significantly generated formaldehyde and formic acid in striatal homogenates. Formaldehyde was the most toxic metabolite to differentiated PC12 pheochromocytoma cells in cell culture studies, indicating that formaldehyde formed endogenously may contribute to neuronal damage in excessive methylation conditions. Subtoxic concentration of formaldehyde decreased the expression of tyrosine hydroxylase, the limiting factor in dopamine synthesis. Formaldehyde was more toxic to catecholaminergic PC12 cells than C6 glioma cells, indicating that neurons are more vulnerable to formaldehyde than glia cells. Significance We suggest that excessive carboxylmethylation of proteins might be involved in the SAM-induced PD-like changes and in the aging process via the toxic effects of formaldehyde. PMID:18930743

  20. Potential for Development of an Escherichia coli—Based Biosensor for Assessing Bioavailable Methionine: A Review

    PubMed Central

    Chalova, Vesela I.; Froelich, Clifford A.; Ricke, Steven C.

    2010-01-01

    Methionine is an essential amino acid for animals and is typically considered one of the first limiting amino acids in animal feed formulations. Methionine deficiency or excess in animal diets can lead to sub-optimal animal performance and increased environmental pollution, which necessitates its accurate quantification and proper dosage in animal rations. Animal bioassays are the current industry standard to quantify methionine bioavailability. However, animal-based assays are not only time consuming, but expensive and are becoming more scrutinized by governmental regulations. In addition, a variety of artifacts can hinder the variability and time efficacy of these assays. Microbiological assays, which are based on a microbial response to external supplementation of a particular nutrient such as methionine, appear to be attractive potential alternatives to the already established standards. They are rapid and inexpensive in vitro assays which are characterized with relatively accurate and consistent estimation of digestible methionine in feeds and feed ingredients. The current review discusses the potential to develop Escherichia coli-based microbial biosensors for methionine bioavailability quantification. Methionine biosynthesis and regulation pathways are overviewed in relation to genetic manipulation required for the generation of a respective methionine auxotroph that could be practical for a routine bioassay. A prospective utilization of Escherichia coli methionine biosensor would allow for inexpensive and rapid methionine quantification and ultimately enable timely assessment of nutritional profiles of feedstuffs. PMID:22319312

  1. Dissimilation of Methionine by a Demethiolase of Aspergillus Species1

    PubMed Central

    Ruiz-Herrera, José; Starkey, Robert L.

    1969-01-01

    Enzyme preparations obtained from the mycelium of Aspergillus species broke down methionine by co-dissimilation. The deaminase and demethiolase activities of crude extracts were increased 100-fold by precipitation with (NH4)2SO4 and column chromatography on diethylaminoethyl cellulose. The enzyme acted on d-methionine but not on l-methionine. The enzyme was labile: it was inactivated by oxygen and ascorbic acid but ethylenediaminetetraacetic acid and mercaptoethanol preserved its activity. Enzyme activity decreased even at 4 and −30 C and was lost rapidly above 45 C. It was most rapid at 35 C and at pH 8.0 to 9.0. For the following reasons, it was concluded that deamination and demethiolation of methionine were effected by the same enzyme: both activities increased equally at each stage of purification; ammonia, methanethiol, and α-keto butyric acid were formed in amounts equivalent to the amount of methionine dissimilated; the Km and optimal pH for formation of both keto acid and methanethiol were the same; both activities remained in the same fractions that were separated by electrophoresis and the activities were equivalent. The purified enzyme demethiolated α-keto methionine and α-hydroxy methionine and split the sulfur linkage of ethionine but did not cleave cystathionine. Few amino acids were deaminated. The enzyme was sensitive to some carbonyl and sulfhydryl reagents and was relatively insensitive to heavy metals other than Hg++. The Km was 1.3 × 10−3 to 1.5 × 10−3m at pH 7.0. No requirement for cofactors was noted, and attempts to dissociate the enzyme, including dialysis with hydroxylamine, were unsuccessful. PMID:5370277

  2. Methionine salvage pathway in relation to ethylene biosynthesis

    SciTech Connect

    Miyazaki, J.H.

    1987-01-01

    The recycling of methionine during ethylene biosynthesis (the methionine cycle) was studied. During ethylene biosynthesis, the H/sub 3/CS-group of S-adenosylmethionine (SAM) is released at 5'-methylthioadenosine (MTA), which is recycled to methionine via 5'-methylthioribose (MTS). In mungbean hypocotyls and cell-free extracts of avocado fruit, (/sup 14/C)MTR was converted to labeled methionine via 2-keto-4-methylthiobutyric acid (KMB) and 2-hydroxy-4-methylthiobutyric acid (HMB) as intermediates. Radioactive tracer studies showed that KMB was converted readily in vivo and in vitro to methionine, while HMB was converted much more slowly. The conversion of KMB to methionine by dialyzed avocado extract required an amino group donor. Among several potential donors tested, L-glutamine was the most efficient. Incubation of (ribose-U-/sup 14/C)MTR with avocado extract resulted in the production of (/sup 14/C)formate, with little evolution of other /sup 14/C-labeled one-carbon compounds, indicating that the conversion of MTR to KMB involves a loss of formate, presumably from C-1 of MTR.

  3. Degradation of Amino Acids and Structure in Model Proteins and Bacteriophage MS2 by Chlorine, Bromine, and Ozone.

    PubMed

    Choe, Jong Kwon; Richards, David H; Wilson, Corey J; Mitch, William A

    2015-11-17

    Proteins are important targets of chemical disinfectants. To improve the understanding of disinfectant-protein reactions, this study characterized the disinfectant:protein molar ratios at which 50% degradation of oxidizable amino acids (i.e., Met, Tyr, Trp, His, Lys) and structure were observed during HOCl, HOBr, and O3 treatment of three well-characterized model proteins and bacteriophage MS2. A critical question is the extent to which the targeting of amino acids is driven by their disinfectant rate constants rather than their geometrical arrangement. Across the model proteins and bacteriophage MS2 (coat protein), differing widely in structure, methionine was preferentially targeted, forming predominantly methionine sulfoxide. This targeting concurs with its high disinfectant rate constants and supports its hypothesized role as a sacrificial antioxidant. Despite higher HOCl and HOBr rate constants with histidine and lysine than for tyrosine, tyrosine generally was degraded in preference to histidine, and to a lesser extent, lysine. These results concur with the prevalence of geometrical motifs featuring histidines or lysines near tyrosines, facilitating histidine and lysine regeneration upon Cl[+1] transfer from their chloramines to tyrosines. Lysine nitrile formation occurred at or above oxidant doses where 3,5-dihalotyrosine products began to degrade. For O3, which lacks a similar oxidant transfer pathway, histidine, tyrosine, and lysine degradation followed their relative O3 rate constants. Except for its low reactivity with lysine, the O3 doses required to degrade amino acids were as low as or lower than for HOCl or HOBr, indicating its oxidative efficiency. Loss of structure did not correlate with loss of particular amino acids, suggesting the need to characterize the oxidation of specific geometric motifs to understand structural degradation.

  4. Factors affecting lysine degradation by ruminal fusobacteria.

    PubMed

    Russell, James B

    2006-04-01

    Fusobacterium necrophorum can readily be enriched from the rumen with lysine, and its deamination rate is very rapid. The addition of F. necrophorum JB2 to mixed ruminal bacteria significantly increased lysine degradation, but only if the ratio of ruminal fluid to basal medium was less than 25%. If more ruminal fluid (pH 6.1) was added, ammonia production decreased by as much as 80%. Clarified, autoclaved ruminal fluid was also inhibitory. When F. necrophorum JB2 was grown in a lysine-limited continuous culture (0.1 h(-1) dilution rate) and pH was decreased using HCl, optical density decreased linearly, and the culture washed out at pH 5.6. Batch cultures of F. necrophorum JB2 deaminated as much lysine at pH 6.1 as at pH 6.6, but only if fermentation acids were not present. Sodium acetate (100 mM) had little effect at pH 6.6, but the same concentration inhibited ammonia production by 80% at pH 6.1. The idea that fermentation acids could prevent the enrichment of fusobacteria in vivo was supported by the observation that dietary lysine supplementation did not enhance the lysine deamination rate of the mixed ruminal bacteria.

  5. Oxidation of Methionine Residues in Polypeptide Ions Via Gas-Phase Ion/Ion Chemistry

    NASA Astrophysics Data System (ADS)

    Pilo, Alice L.; McLuckey, Scott A.

    2014-06-01

    The gas-phase oxidation of methionine residues is demonstrated here using ion/ion reactions with periodate anions. Periodate anions are observed to attach in varying degrees to all polypeptide ions irrespective of amino acid composition. Direct proton transfer yielding a charge-reduced peptide ion is also observed. In the case of methionine and, to a much lesser degree, tryptophan-containing peptide ions, collisional activation of the complex ion generated by periodate attachment yields an oxidized peptide product (i.e., [M + H + O]+), in addition to periodic acid detachment. Detachment of periodic acid takes place exclusively for peptides that do not contain either a methionine or tryptophan side chain. In the case of methionine-containing peptides, the [M + H + O]+ product is observed at a much greater abundance than the proton transfer product (viz., [M + H]+). Collisional activation of oxidized Met-containing peptides yields a signature loss of 64 Da from the precursor and/or product ions. This unique loss corresponds to the ejection of methanesulfenic acid from the oxidized methionine side chain and is commonly used in solution-phase proteomics studies to determine the presence of oxidized methionine residues. The present work shows that periodate anions can be used to `label' methionine residues in polypeptides in the gas phase. The selectivity of the periodate anion for the methionine side chain suggests several applications including identification and location of methionine residues in sequencing applications.

  6. Oxidation of methionine residues in polypeptide ions via gas-phase ion/ion chemistry.

    PubMed

    Pilo, Alice L; McLuckey, Scott A

    2014-06-01

    The gas-phase oxidation of methionine residues is demonstrated here using ion/ion reactions with periodate anions. Periodate anions are observed to attach in varying degrees to all polypeptide ions irrespective of amino acid composition. Direct proton transfer yielding a charge-reduced peptide ion is also observed. In the case of methionine and, to a much lesser degree, tryptophan-containing peptide ions, collisional activation of the complex ion generated by periodate attachment yields an oxidized peptide product (i.e., [M + H + O](+)), in addition to periodic acid detachment. Detachment of periodic acid takes place exclusively for peptides that do not contain either a methionine or tryptophan side chain. In the case of methionine-containing peptides, the [M + H + O](+) product is observed at a much greater abundance than the proton transfer product (viz., [M + H](+)). Collisional activation of oxidized Met-containing peptides yields a signature loss of 64 Da from the precursor and/or product ions. This unique loss corresponds to the ejection of methanesulfenic acid from the oxidized methionine side chain and is commonly used in solution-phase proteomics studies to determine the presence of oxidized methionine residues. The present work shows that periodate anions can be used to 'label' methionine residues in polypeptides in the gas phase. The selectivity of the periodate anion for the methionine side chain suggests several applications including identification and location of methionine residues in sequencing applications.

  7. Acid-base interactions and secondary structures of poly-L-lysine probed by 15N and 13C solid state NMR and Ab initio model calculations.

    PubMed

    Dos, Alexandra; Schimming, Volkmar; Tosoni, Sergio; Limbach, Hans-Heinrich

    2008-12-11

    The interactions of the 15N-labeled amino groups of dry solid poly-L-lysine (PLL) with various halogen and oxygen acids HX and the relation to the secondary structure have been studied using solid-state 15N and 13C CPMAS NMR spectroscopy (CP = cross polarization and MAS = magic angle spinning). For comparison, 15N NMR spectra of an aqueous solution of PLL were measured as a function of pH. In order to understand the effects of protonation and hydration on the 15N chemical shifts of the amino groups, DFT and chemical shielding calculations were performed on isolated methylamine-acid complexes and on periodic halide clusters of the type (CH3NH3(+)X(-))n. The combined experimental and computational results reveal low-field shifts of the amino nitrogens upon interaction with the oxygen acids HX = HF, H2SO4, CH3COOH, (CH3)2POOH, H3PO4, HNO3, and internal carbamic acid formed by reaction of the amino groups with gaseous CO2. Evidence is obtained that only hydrogen-bonded species of the type (Lys-NH2***H-X)n are formed in the absence of water. 15N chemical shifts are maximum when H is located in the hydrogen bond center and then decrease again upon full protonation, as found for aqueous solution at low pH. By contrast, halogen acids interact in a different way. They form internal salts of the type (Lys-NH3(+)X(-))n via the interaction of many acid-base pairs. This salt formation is possible only in the beta-sheet conformation. By contrast, the formation of hydrogen-bonded complexes can occur both in beta-sheet domains as well as in alpha-helical domains. The 15N chemical shifts of the protonated ammonium groups increase when the size of the interacting halogen anions is increased from chloride to iodide and when the number of the interacting anions is increased. Thus, the observed high-field 15N shift of ammonium groups upon hydration is the consequence of replacing interacting halogen atoms by oxygen atoms.

  8. Evidence in support of lysine 77 and histidine 96 as acid-base catalytic residues in saccharopine dehydrogenase from Saccharomyces cerevisiae.

    PubMed

    Kumar, Vidya Prasanna; Thomas, Leonard M; Bobyk, Kostyantyn D; Andi, Babak; Cook, Paul F; West, Ann H

    2012-01-31

    Saccharopine dehydrogenase (SDH) catalyzes the final reaction in the α-aminoadipate pathway, the conversion of l-saccharopine to l-lysine (Lys) and α-ketoglutarate (α-kg) using NAD⁺ as an oxidant. The enzyme utilizes a general acid-base mechanism to conduct its reaction with a base proposed to accept a proton from the secondary amine of saccharopine in the oxidation step and a group proposed to activate water to hydrolyze the resulting imine. Crystal structures of an open apo form and a closed form of the enzyme with saccharopine and NADH bound have been determined at 2.0 and 2.2 Å resolution, respectively. In the ternary complex, a significant movement of domain I relative to domain II that closes the active site cleft between the two domains and brings H96 and K77 into the proximity of the substrate binding site is observed. The hydride transfer distance is 3.6 Å, and the side chains of H96 and K77 are properly positioned to act as acid-base catalysts. Preparation of the K77M and H96Q single-mutant and K77M/H96Q double-mutant enzymes provides data consistent with their role as the general acid-base catalysts in the SDH reaction. The side chain of K77 initially accepts a proton from the ε-amine of the substrate Lys and eventually donates it to the imino nitrogen as it is reduced to a secondary amine in the hydride transfer step, and H96 protonates the carbonyl oxygen as the carbinolamine is formed. The K77M, H976Q, and K77M/H96Q mutant enzymes give 145-, 28-, and 700-fold decreases in V/E(t) and >10³-fold increases in V₂/K(Lys)E(t) and V₂/K(α-kg)E(t) (the double mutation gives >10⁵-fold decreases in the second-order rate constants). In addition, the K77M mutant enzyme exhibits a primary deuterium kinetic isotope effect of 2.0 and an inverse solvent deuterium isotope effect of 0.77 on V₂/K(Lys). A value of 2.0 was also observed for (D)(V₂/K(Lys))(D₂O) when the primary deuterium kinetic isotope effect was repeated in D₂O, consistent with a

  9. Effects of calcium salts of long-chain fatty acids and rumen-protected methionine on plasma concentrations of ghrelin, glucagon-like peptide-1 (7 to 36) amide and pancreatic hormones in lactating cows.

    PubMed

    Fukumori, R; Sugino, T; Shingu, H; Moriya, N; Hasegawa, Y; Kojima, M; Kangawa, K; Obitsu, T; Kushibiki, S; Taniguchi, K

    2012-02-01

    Our objective was to determine the effects of calcium salts of long-chain fatty acids (CLFAs) and rumen-protected methionine (RPM) on plasma concentrations of ghrelin, glucagon-like peptide-1 (7 to 36) amide, and pancreatic hormones in lactating cows. Four Holstein cows in midlactation were used in a 4 by 4 Latin square experiment in each 2-wk period. Cows were fed corn silage-based diets with supplements of CLFAs (1.5% added on dry matter basis), RPM (20 g/d), CLFAs plus RPM, and without supplement. Jugular blood samples were taken from 1 h before to 2 h after morning feeding at 10-min intervals on day 12 of each period. CLFAs decreased dry matter intake, but RPM did not affect dry matter intake. Both supplements of CLFAs and RPM did not affect metabolizable energy intake and milk yield and composition. Plasma concentrations of NEFAs, triglyceride (TG), and total cholesterol (T-Cho) were increased with CLFAs alone, but increases of plasma concentrations of TG and T-Cho were moderated by CLFAs plus RPM. Calcium salts of long-chain fatty acids increased plasma ghrelin concentration, and the ghrelin concentration with CLFAs plus RPM was the highest among the treatments. Plasma concentrations of glucagon-like peptide-1, glucagon, and insulin were decreased with CLFAs, whereas adding RPM moderated the decrease of plasma glucagon concentration by CLFAs. These results indicate that the addition of methionine to cows given CLFAs increases plasma concentrations of ghrelin and glucagon associated with the decrease in plasma concentrations of TG and T-Cho.

  10. Effects of zinc and sodium monensin on ruminal degradation of lysine-HCl and liquid 2-hydroxy-4-methylthiobutanoic acid.

    PubMed

    Bateman, H G; Williams, C C; Gantt, D T; Chung, Y H; Beem, A E; Stanley, C C; Goodier, G E; Hoyt, P G; Ward, J D; Bunting, L D

    2004-08-01

    Four nonlactating, mature, Holstein cows were fitted with ruminal cannula and used in a 4 x 4 Latin square-designed experiment to evaluate the impact of supplemental Zn and monensin on ruminal degradation of Lys and liquid 2-hydroxy-4-methylthiobutanoic acid (HMB). Cows were fed 4.54 kg (as fed) of alfalfa hay top-dressed with 4.54 kg (as fed) concentrate once daily. Concentrates were formulated to provide 0 or 500 mg/kg of Zn as ZnSO4 and 0 or 40 mg/kg of monensin in the total diet. Zinc supplementation provided approximately 22-fold greater dietary Zn than estimated by NRC requirements. On d 14 of each period, cows were dosed via the rumen cannula with 50 g of HMB and 100 g of Lys-HCl, and the concentrations of Lys and HMB were monitored every 0.5 h for 8 h. Supplemental Zn tended to decrease the proportion of acetate in ruminal fluid postfeeding and increased the proportion of propionate in ruminal fluid postfeeding. Supplemental Zn increased mean fluid passage rate from the rumen. Monensin decreased the proportion of acetate and increased the mean proportion of propionate in ruminal fluid, resulting in a decrease in the ratio of acetate to propionate. Monensin also increased the mean fluid passage rate from the rumen. Neither Zn nor monensin affected the apparent rate of ruminal disappearance of HMB or Lys. However, Zn and monensin interacted to alter the ruminal degradability of free Lys but not HMB. These data indicate that Zn and monensin may interact to alter ruminal degradability of free amino acids.

  11. Diversity of L-methionine catabolism pathways in cheese-ripening bacteria.

    PubMed

    Bonnarme, P; Psoni, L; Spinnler, H E

    2000-12-01

    Enzymatic activities that could be involved in methanethiol generation in five cheese-ripening bacteria were assayed, and the major sulfur compounds produced were identified. L-Methionine and alpha-keto-gamma-methyl-thio-butyric acid demethiolating activities were detected in whole cells and cell extracts (CFEs) of all the bacteria tested. No L-methionine deaminase activity could be detected in any of the ripening bacteria and L-methionine aminotransferase was detected in CFEs of Brevibacterium linens, Micrococcus luteus, and Corynebacterium glutamicum. The results suggest that several pathways for L-methionine catabolism probably coexist in these ripening bacteria.

  12. The low-methionine content of vegan diets may make methionine restriction feasible as a life extension strategy.

    PubMed

    McCarty, Mark F; Barroso-Aranda, Jorge; Contreras, Francisco

    2009-02-01

    Recent studies confirm that dietary methionine restriction increases both mean and maximal lifespan in rats and mice, achieving "aging retardant" effects very similar to those of caloric restriction, including a suppression of mitochondrial superoxide generation. Although voluntary caloric restriction is never likely to gain much popularity as a pro-longevity strategy for humans, it may be more feasible to achieve moderate methionine restriction, in light of the fact that vegan diets tend to be relatively low in this amino acid. Plant proteins - especially those derived from legumes or nuts - tend to be lower in methionine than animal proteins. Furthermore, the total protein content of vegan diets, as a function of calorie content, tends to be lower than that of omnivore diets, and plant protein has somewhat lower bioavailability than animal protein. Whole-food vegan diets that moderate bean and soy intake, while including ample amounts of fruit and wine or beer, can be quite low in methionine, while supplying abundant nutrition for health (assuming concurrent B12 supplementation). Furthermore, low-fat vegan diets, coupled with exercise training, can be expected to promote longevity by decreasing systemic levels of insulin and free IGF-I; the latter effect would be amplified by methionine restriction - though it is not clear whether IGF-I down-regulation is the sole basis for the impact of low-methionine diets on longevity in rodents.

  13. Thyroid metabolism in the recessive sex-linked dwarf female chicken. 4. The influence of exogenous thyroid hormones on amino acid uptake by plasma and tissues.

    PubMed

    Grandhi, R R; Brown, R G; Reinhart, B S; Summers, J D

    1975-03-01

    The influence of exogenous triiodothyronine (T3) or tetraiodothyronine (T4) on the incorporation of 3H-labelled methionine, alanine and lysine into plasma, liver and kidney was studied in 4 wk. old dwarf and non-dwarf female, White Leghorn chickens. The response to exogenous T3 or T4 of the birds was directly dependent on the dwarf status as well as the tissue and/or amino acid studied. In general, there was a decreased amino acid uptake by dwarfs and T3 and/or T4 depressed amino acid uptake in all combinations studied except for the uptake of lysine by the kidney of the dwarf. In that tissue, T4 administration caused a significant increase in lysine incorporation. The results found probably were due to different tissue amino compositions or rates of synthesis of proteins by dwarf birds when compared to normals and a differential sensitivity of dwarf birds to changed T3/T4 ratios.

  14. Effect of L-lysine on expression of selected genes, serum concentration of amino acids, muscle growth and performance of growing pigs.

    PubMed

    Morales, A; García, H; Arce, N; Cota, M; Zijlstra, R T; Araiza, B A; Cervantes, M

    2015-08-01

    Lysine (Lys) is the first limiting amino acid (AA) in most feed formulations for pigs and most abundant, along with leucine, in muscle proteins. An experiment was conducted with 17 pigs (17.7 ± 0.05 kg initial BW) to identify a role of dietary Lys in the control of protein synthesis in pigs. Fourteen pigs were randomly assigned to one of the two wheat-based dietary treatments: Lys-deficient, 3.0 g/kg (DEF) and Lys-adequate, 10.8 g/kg (ADE). Samples from jejunum mucosa, liver, Longissumus and Semitendinosus muscles, and blood were collected. The other three pigs were sacrificed at the beginning of the trial to measure basal carcass composition. Weight gain, gain:feed ratio, Lys intake and loin eye area were greater in ADE than in DEF pigs (p < 0.01). Muscle-related carcass characteristics were better, and myosin heavy chain IIb expression (MyHC IIb) in Semitendinosus was higher in ADE than in DEF pigs. Expression of AA transporters CAT-1 was lower (p < 0.05), serum Lys was higher and serum Val was lower in pigs fed the ADE diet. The higher muscularity, MyHC IIb expression in Semitendinosus muscle and Lys serum of pigs fed the ADE diet suggest that Lys increases growth rate not only by functioning as protein construction unit but also as potential control of the protein synthesis process.

  15. Probing the role of lysine 16 in ras p[sup 21] protein with unnatural amino acids

    SciTech Connect

    Chung, H.H.; Benson, D.R.; Schultz, P.G. )

    1993-07-14

    Mamalian proteins encoded by the ras gene are thought to function as regulators of various signal transduction processes involved in cell growth and differentiation. The chemical basis for the regulation is cycling of the protein between the inactive guanosine diphosphate (GDP)-bound state and the active guanosine triphophate (GTP)-bound state. Loop 1 of ras contains the GXXXXGK(S/T) motif (residues 10-17), which is found in all ras-related proteins, G proteins, and other nucleotide-binding proteins. Structural and biochemical studies have suggested that Lys 16 of loop 1 is critical for substrate binding and catalysis. The [epsilon]-amino group is involved in ion pair interactions with the [Beta]-and [gamma]-phosphates of GTP and forms hydrogen bonds with the main-chain oxygens of Gly 10 and Ala 11. In order to better understand the role of the key residue in ras function, we have replaced Lys 16 with a number of unnatural amino acid analogues, including (aminoethyl)cysteine, (hydroxyethyl)cysteine, (aminoethyl)homocysteine, and ornithine. We were surprised to find that the [open quotes]unnatural[close quotes] mutant ras proteins retained high levels of GAP-stimulated GTPase activity and GTP dissociation rates comparable to that of wild-type ras. These results may indicate that, in the GAP-activated form, Lys 16 is not involved in transition-state stabilization or GTP binding. However, replacement of Lys 16 with the isosteric uncharged Lys analogue, (hydroxyethyl)cysteine, led to a complete loss of GAP-stimulated GTPase activity, demonstrating the importance of the charged ammonium side chain. 22 refs., 1 fig., 1 tab.

  16. Accumulation, selection and covariation of amino acids in sieve tube sap of tansy (Tanacetum vulgare) and castor bean (Ricinus communis): evidence for the function of a basic amino acid transporter and the absence of a γ-amino butyric acid transporter.

    PubMed

    Bauer, Susanne N; Nowak, Heike; Keller, Frank; Kallarackal, Jose; Hajirezaei, Mohamad-Reza; Komor, Ewald

    2014-09-01

    Sieve tube sap was obtained from Tanacetum by aphid stylectomy and from Ricinus after apical bud decapitation. The amino acids in sieve tube sap were analyzed and compared with those from leaves. Arginine and lysine accumulated in the sieve tube sap of Tanacetum more than 10-fold compared to the leaf extracts and they were, together with asparagine and serine, preferably selected into the sieve tube sap, whereas glycine, methionine/tryptophan and γ-amino butyric acid were partially or completely excluded. The two basic amino acids also showed a close covariation in sieve tube sap. The acidic amino acids also grouped together, but antagonistic to the other amino acids. The accumulation ratios between sieve tube sap and leaf extracts were smaller in Ricinus than in Tanacetum. Arginine, histidine, lysine and glutamine were enriched and preferentially loaded into the phloem, together with isoleucine and valine. In contrast, glycine and methionine/tryptophan were partially and γ-amino butyric acid almost completely excluded from sieve tube sap. The covariation analysis grouped arginine together with several neutral amino acids. The acidic amino acids were loaded under competition with neutral amino acids. It is concluded from comparison with the substrate specificities of already characterized plant amino acid transporters, that an AtCAT1-like transporter functions in phloem loading of basic amino acids, whereas a transporter like AtGAT1 is absent in phloem. Although Tanacetum and Ricinus have different minor vein architecture, their phloem loading specificities for amino acids are relatively similar.

  17. Amino acid and energy interrelationships in growing beef steers: II. Effects of energy intake and metabolizable lysine supply on growth.

    PubMed

    Ludden, P A; Kerley, M S

    1998-12-01

    We conducted three experiments to determine the optimal metabolizable Lys:net energy ratio for growth of beef calves. The single basal diet fed contained corn (56.1%), soybean hulls (18%), cottonseed hulls (15%), animal fat (4.25%), and corn gluten meal (5.6%). In Exp. 1, 54 steers were individually fed the basal diet at 1.5, 2.25, and 3.0 times NEm requirement; rations were top-dressed with 3.4 g of rumen-stable (RS) Met and either 0, 2, 4, 6, 8, or 12 g of RS-Lys daily. An additional 18 steers were fed the same three levels of energy and supplemented with 125 g of blood meal per steer. In Exp. 2, 68 crossbred steers were subjected to the same experimental protocol, with the exception that only the two highest levels of energy were used. Of these steers, 48 were fed individually and received the RS-Lys treatments; the remaining 20 steers received 125 g of blood meal per steer. No interaction (P > .10) was detected between level of supplemental Lys and energy intake in Exp. 1 or 2. Supplementation with RS-Lys improved (P < .01) ADG in Exp. 1, but it had no effect (P > .10) on growth in Exp. 2. The Lys requirement estimates were 44.3 and 51.3 g/d, corresponding to maximal growth rates of 1.21 and 1.64 kg/d for the 2.25 and 3.0 times maintenance treatments, respectively. Comparing the growth rates of steers fed supplemental Lys with those of steers fed blood meal in Exp. 1 and 2 revealed an ADG advantage (P < .03) with blood meal supplementation. To confirm the blood meal response, Exp. 3 used 75 crossbred steers fed the basal diet at 3.0 times NEm requirement plus either 3.4 g RS-Met, 3.4 g RS-Met and 12 g RS-Lys, or 125 g of blood meal per steer. Blood meal supplementation improved (P < .01) growth of steers over those fed supplemental Met or Met plus Lys. Although a distinct relationship between amino acid requirements and energy supply may exist, Lys and Met were not first-limiting in these experiments, or selective supplementation with undegradable protein may

  18. Serine Metabolism Supports the Methionine Cycle and DNA/RNA Methylation through De Novo ATP Synthesis in Cancer Cells

    PubMed Central

    Maddocks, Oliver D.K.; Labuschagne, Christiaan F.; Adams, Peter D.; Vousden, Karen H.

    2016-01-01

    Summary Crosstalk between cellular metabolism and the epigenome regulates epigenetic and metabolic homeostasis and normal cell behavior. Changes in cancer cell metabolism can directly impact epigenetic regulation and promote transformation. Here we analyzed the contribution of methionine and serine metabolism to methylation of DNA and RNA. Serine can contribute to this pathway by providing one-carbon units to regenerate methionine from homocysteine. While we observed this contribution under methionine-depleted conditions, unexpectedly, we found that serine supported the methionine cycle in the presence and absence of methionine through de novo ATP synthesis. Serine starvation increased the methionine/S-adenosyl methionine ratio, decreasing the transfer of methyl groups to DNA and RNA. While serine starvation dramatically decreased ATP levels, this was accompanied by lower AMP and did not activate AMPK. This work highlights the difference between ATP turnover and new ATP synthesis and defines a vital function of nucleotide synthesis beyond making nucleic acids. PMID:26774282

  19. [The microflora of sourdough. XVIII. The protein degrading capabilities of lactic acid bacteria in sourdough].

    PubMed

    Spicher, G; Nierle, W

    1984-05-01

    Acidification of the dough by the use of sourdough or acidifiers is necessary not only for good baking quality of rye flour but it is also very important for development of the typical sensory characteristics of rye bread. We confirmed that the lactic acid bacteria of sour dough are proteolytic. Proteolytic effects are observed in the increase of the amino acid content during fermentation. A marked increase was found in the content of leucine, alanine, valine, isoleucine, glutamic acid, glutamine, arginine, lysine, methionine, phenylalanine, tyrosine and serine. Lactobacillus plantarum showed a higher proteolytic activity than L. brevis ssp. lindneri or L. fructivorans.

  20. Synthesis of Lysine Methyltransferase Inhibitors

    NASA Astrophysics Data System (ADS)

    Ye, Tao; Hui, Chunngai

    2015-07-01

    Lysine methyltransferase which catalyze methylation of histone and nonhistone proteins, play a crucial role in diverse biological processes and has emerged as a promising target for the development of various human diseases, including cancer, inflammation, and psychiatric disorders. However, inhibiting Lysine methyltransferases selectively has presented many challenges to medicinal chemists. During the past decade, lysine methyltransferase inhibitors covering many different structural classes have been designed and developed. In this review, we describe the development of selective, small-molecule inhibitors of lysine methyltransferases with an emphasis on their discovery and chemical synthesis. We highlight the current state of lysine methyltransferase inhibitors and discuss future directions and opportunities for lysine methyltransferase inhibitor discovery.

  1. Methionine Metabolism Alters Oxidative Stress Resistance via the Pentose Phosphate Pathway.

    PubMed

    Campbell, Kate; Vowinckel, Jakob; Keller, Markus A; Ralser, Markus

    2016-04-01

    Nutrient uptake and metabolism have a significant impact on the way cells respond to stress. The amino acid methionine is, in particular, a key player in the oxidative stress response, and acting as a reactive oxygen species scavenger, methionine is implicated in caloric restriction phenotypes and aging. We here provide evidence that some effects of methionine in stress situations are indirect and caused by altered activity of the nicotinamide adenine dinucleotide phosphate (NADPH) producing oxidative part of the pentose phosphate pathway (PPP). In Saccharomyces cerevisiae, both methionine prototrophic (MET15) and auxotrophic (met15Δ) cells supplemented with methionine showed an increase in PPP metabolite concentrations downstream of the NADPH producing enzyme, 6-phosphogluconate dehydrogenase. Proteomics revealed this enzyme to also increase in expression compared to methionine self-synthesizing cells. Oxidant tolerance was increased in cells preincubated with methionine; however, this effect was abolished when flux through the oxidative PPP was prevented by deletion of its rate limiting enzyme, ZWF1. Stress resistance phenotypes that follow methionine supplementation hence involve the oxidative PPP. Effects of methionine on oxidative metabolism, stress signaling, and aging have thus to be seen in the context of an altered activity of this NADP reducing pathway.

  2. Response of growing goslings to dietary supplementation with methionine and betaine.

    PubMed

    Yang, Z; Wang, Z Y; Yang, H M; Zhao, F Z; Kong, L L

    2016-12-01

    An experiment with a 2 × 3 factorial design with two concentrations of dietary betaine (0 and 600 mg/kg) and three dietary concentrations of methionine (0, 600 and 1200 mg/kg) was conducted using goslings to estimate growth, nutrient utilisation and digestibility of amino acids from 21 to 70 d of age. Three hundred geese were randomised at 18 d of age into 6 groups with 5 replicates per treatment and 10 geese per replicate. Increasing dietary concentrations of methionine gave a linear increase in body weight and average daily gain. The coefficient of crude fat retention increased as dietary methionine increased and there was a significant non-linear response to increasing dietary methionine. Similarly, increasing supplemental methionine gave linear increases in the digestibility of methionine and cysteine. The results of this study indicated that optimal dietary supplementation of methionine could increase growth performance and methionine and cysteine utilisation in growing goslings. Betaine supplementation had no apparent sparing effect on methionine needs for growth performance, but did improve the apparent cysteine digestibility.

  3. Methionine Metabolism Alters Oxidative Stress Resistance via the Pentose Phosphate Pathway

    PubMed Central

    Campbell, Kate; Vowinckel, Jakob; Keller, Markus A.

    2016-01-01

    Abstract Nutrient uptake and metabolism have a significant impact on the way cells respond to stress. The amino acid methionine is, in particular, a key player in the oxidative stress response, and acting as a reactive oxygen species scavenger, methionine is implicated in caloric restriction phenotypes and aging. We here provide evidence that some effects of methionine in stress situations are indirect and caused by altered activity of the nicotinamide adenine dinucleotide phosphate (NADPH) producing oxidative part of the pentose phosphate pathway (PPP). In Saccharomyces cerevisiae, both methionine prototrophic (MET15) and auxotrophic (met15Δ) cells supplemented with methionine showed an increase in PPP metabolite concentrations downstream of the NADPH producing enzyme, 6-phosphogluconate dehydrogenase. Proteomics revealed this enzyme to also increase in expression compared to methionine self-synthesizing cells. Oxidant tolerance was increased in cells preincubated with methionine; however, this effect was abolished when flux through the oxidative PPP was prevented by deletion of its rate limiting enzyme, ZWF1. Stress resistance phenotypes that follow methionine supplementation hence involve the oxidative PPP. Effects of methionine on oxidative metabolism, stress signaling, and aging have thus to be seen in the context of an altered activity of this NADP reducing pathway. Antioxid. Redox Signal. 24, 543–547. PMID:26596469

  4. Optimization of a gas chromatography-mass spectrometry method with methyl chloroformate derivatization for quantification of amino acids in plant tissue.

    PubMed

    Vancompernolle, Bram; Croes, Kim; Angenon, Geert

    2016-04-01

    Rapid, easy and reliable quantification of amino acids is crucial in research on plant amino acid metabolism and nutritional improvement of crops via enrichment of essential amino acids. A recently reported analysis method, based on solid phase extraction (SPE), derivatization with methyl chloroformate and gas chromatography-mass spectrometry was optimized and tested on three-week-old Arabidopsis thaliana leaf tissues. Optimization of the SPE cleanup yielded recovery rates of minimum 95% for all amino acids (except arginine). Variations in accuracy and precision did not exceed 12.5%, except for cysteine, histidine and tryptophane, which were excluded from analysis. Quantification of overlapping peaks for isoleucine/threonine and proline/asparagine was possible by selection of two specific fragment ions for each amino acid. Of the 16 selected amino acids, 14 were quantified successfully in at least 75% of the samples, while methionine and tyrosine were only quantifiable in 6% and 42%, respectively. A case study on the aspartate super pathway confirmed the applicability of the optimized method on wild type and genetically modified plants: external supplementation of methionine or lysine yielded a 146-fold or 27-fold increase in the respective absolute amino acid levels compared with the control treatment. Induced expression of dhdps-r1 (a mutated lysine biosynthesis gene encoding a feedback insensitive enzyme) caused an 83-fold increase in absolute lysine levels.

  5. Methionine splanchnic uptake is increased in critically ill children

    Technology Transfer Automated Retrieval System (TEKTRAN)

    During critical illness the splanchnic area is profoundly affected. There is no information on splanchnic uptake of amino acids in vivo, in critically ill children. Methionine splanchnic uptake in critically ill children will differ from estimates in healthy adults. We studied 24 critically ill chil...

  6. Simple method for the simultaneous analysis of pipecolic acid and lysine by high-performance liquid chromatography and its application to rumen liquor and plasma of ruminants.

    PubMed

    Hussain-Yusuf, H; Onodera, R; Nasser, M E; Sato, H

    1999-11-26

    A high-performance liquid chromatography method for the simultaneous determination of pipecolic acid (Pip) and lysine (Lys), a precursor of Pip, in the rumen liquor and plasma of ruminant animals was established. Samples of rumen liquor and plasma were deproteinized with 50% acetonitrile and derivatized with a fluorescent agent 9-fluorenylmethyloxy carbonyl chloride (Fmoc-Cl). Chromatographic separation was achieved on a TSK gel ODS-80TM column using a reversed-phase gradient elution system. For the gradient elution, two mobile phases, A and B, were needed, both commonly consisted of: 5 mM L-proline, 2.5 mM cupric sulfate and 6.5 mM ammonium acetate. Mobile phase B additionally contains 50% (v/v) acetonitrile. The pH of both mobile phases was adjusted to 7.0. Derivatized Pip and Lys were detected on a fluorescent detector at excitation and emission wavelengths of 260 and 313 nm, respectively. The calibration curves were linear within the range 0 to 1 mM (r>0.999). The average recoveries for Pip and Lys were 95.9+/-1.8 and 93.2+/-2.5% in rumen liquor and 98.3+/-1.4 and 97.5+/-1.3% in plasma, respectively. The limits of detection for Pip and Lys were 0.6 and 0.7 microM in rumen liquor and 0.01 and 0.05 microM in plasma. The assay has acceptable precision, relative standard deviation (RSD) for reproducibility (within-day and day-to-day variation) were less than 5.2% for aqueous (5.0 microM Pip and Lys), MB9 (5.0 microM Pip and Lys), plasma (7.1 microM Pip and 85.6 microM Lys) and rumen liquor (28.4 microM Pip and 10.2 microM Lys) samples. The levels of Pip and Lys in faunated goats, determined from three animals over a period of two days sampling, were found to be 36.8+/-18.1 and 14.6+/-2.8 microM in rumen liquor, and 7.3+/-2.5 and 137.3+/-38.0 microM in plasma at 1 h after feeding. This is the first report on the normal levels of Pip in the rumen liquor and plasma of faunated goat.

  7. Regulation of lysine biosynthesis and transport genes in bacteria: yet another RNA riboswitch?

    PubMed

    Rodionov, Dmitry A; Vitreschak, Alexey G; Mironov, Andrey A; Gelfand, Mikhail S

    2003-12-01

    Comparative analysis of genes, operons and regulatory elements was applied to the lysine biosynthetic pathway in available bacterial genomes. We report identification of a lysine-specific RNA element, named the LYS element, in the regulatory regions of bacterial genes involved in biosynthesis and transport of lysine. Similarly to the previously described RNA regulatory elements for three vitamins (riboflavin, thiamin and cobalamin), purine and methionine regulons, this regulatory RNA structure is highly conserved on the sequence and structural levels. The LYS element includes regions of lysine-constitutive mutations previously identified in Escherichia coli and Bacillus subtilis. A possible mechanism of the lysine-specific riboswitch is similar to the previously defined mechanisms for the other metabolite-specific riboswitches and involves either transcriptional or translational attenuation in various groups of bacteria. Identification of LYS elements in Gram-negative gamma-proteobacteria, Gram-positive bacteria from the Bacillus/Clostridium group, and Thermotogales resulted in description of the previously uncharacterized lysine regulon in these bacterial species. Positional analysis of LYS elements led to identification of a number of new candidate lysine transporters, namely LysW, YvsH and LysXY. Finally, the most likely candidates for genes of lysine biosynthesis missing in Gram- positive bacteria were identified using the genome context analysis.

  8. From yeast to human: exploring the comparative biology of methionine restriction in extending eukaryotic life span.

    PubMed

    McIsaac, R Scott; Lewis, Kaitlyn N; Gibney, Patrick A; Buffenstein, Rochelle

    2016-01-01

    Methionine restriction is a widely reported intervention for increasing life span in several model organisms. Low circulating levels of methionine are evident in the long-lived naked mole-rat, suggesting that it naturally presents with a life-extending phenotype akin to that observed in methionine-restricted animals. Similarly, long-lived dwarf mice also appear to have altered methionine metabolism. The mechanisms underlying methionine-restriction effects on life-span extension, however, remain unknown, as do their potential connections with caloric restriction, another well-established intervention for prolonging life span. Paradoxically, methionine is enriched in proteins expressed in mitochondria and may itself serve an important role in the detoxification of reactive oxygen species and may thereby contribute to delayed aging. Collectively, we highlight the evidence that modulation of the methionine metabolic network can extend life span-from yeast to humans-and explore the evidence that sulfur amino acids and the concomitant transsulfuration pathway play a privileged role in this regard. However, systematic studies in single organisms (particularly those that exhibit extreme longevity) are still required to distinguish the fundamental principles concerning the role of methionine and other amino acids in regulating life span.

  9. Influence of dietary methionine on the metabolism of selenomethionine in rats

    SciTech Connect

    Butler, J.A.; Beilstein, M.A.; Whanger, P.D. )

    1989-07-01

    To determine the influence of methionine on selenomethionine (SeMet) metabolism, weanling male rats were fed for 8 wk a basal diet marginally deficient in sulfur amino acids, containing 2.0 micrograms selenium (Se)/g as DL-SeMet and supplemented with 0, 0.3, 0.6 or 1.2% DL-methionine. Increased dietary methionine caused decreased selenium deposition in all tissues examined but increased glutathione peroxidase activity in testes, liver and lungs. A positive correlation was found between dietary methionine and the calculated percentage of selenium associated with GSHPx. In a second experiment, {sup 75}SeMet was injected into weanling male rats which had been fed the basal diet containing 2.0 micrograms selenium as DL-SeMet with or without the addition of 1.0% methionine. The selenoamino acid content of tissues and the distribution of {sup 75}Se in erythrocyte proteins were determined. In comparison to the rats fed the basal diet without added methionine, significantly more {sup 75}Se-selenocysteine was found in liver and muscle, more {sup 75}Se was found in erythrocyte GSHPx and less {sup 75}Se was found in erythrocyte hemoglobin of rats fed 1.0% methionine. These data suggest that methionine diverts SeMet from incorporation into general proteins and enhances its conversion to selenocysteine for specific selenium-requiring proteins, such as GSHPx.

  10. Comparative value of L-, and D-methionine supplementation of an oat-based diet for humans.

    PubMed

    Kies, C; Fox, H; Aprahamian, S

    1975-07-01

    Total sulfur-containing amino acids have been found to be the first limiting amino acid in several foods in comparison with human amino acid requirements. Addition of methionine in appropriate amounts to these foods might be expected to improve protein value. Economically, DL-methionine would be preferable to L-methionine for this purpose. However, the comparative tuilization of L- DL-, and D-methionine is unclear. The objective of the current project was to compare the effectiveness of L-, DL-, and D-methionine supplementation of diets based on a food product known to be low in methionine value for human subjects. "Instant" oatmeal was fed to adult subjects to provide 4.0 g of nitrogen/day. In randomly arranged periods, these diets were supplemented with L-, DL-, or D-methionine at two levels (0.58 and 1.16 g of methionine/day). An unsupplemented diet was used in a control period. Diets were adequate in vitamins, minerals, and energy. Mean nitrogen balances of subjects while receiving the L-methionine supplements at the 0.58 and 1.16 g levels were minus0.10 and +0.06 g of nitrogen, respectively. At similar levels of DL-methionine supplementation, nitrogen balances were minus0.12 and minus0.15 g of nitrogen, respectively, and minus0.24 and minus0.18 g of nitrogen with D-methionine supplementation. The mean nitrogen balance when no supplement was used was minus0.22 g of nitrogen. Thus, D-methionine is seemingly poorly utilized by the human. Urinary methionine excretion data supported these results.

  11. Amino Acid Uptake in Arbuscular Mycorrhizal Plants

    PubMed Central

    Whiteside, Matthew D.; Garcia, Maria O.; Treseder, Kathleen K.

    2012-01-01

    We examined the extent to which arbuscular mycorrhizal (AM) fungi root improved the acquisition of simple organic nitrogen (ON) compounds by their host plants. In a greenhouse-based study, we used quantum dots (fluorescent nanoparticles) to assess uptake of each of the 20 proteinaceous amino acids by AM-colonized versus uncolonized plants. We found that AM colonization increased uptake of phenylalanine, lysine, asparagine, arginine, histidine, methionine, tryptophan, and cysteine; and reduced uptake of aspartic acid. Arbuscular mycorrhizal colonization had the greatest effect on uptake of amino acids that are relatively rare in proteins. In addition, AM fungi facilitated uptake of neutral and positively-charged amino acids more than negatively-charged amino acids. Overall, the AM fungi used in this study appeared to improve access by plants to a number of amino acids, but not necessarily those that are common or negatively-charged. PMID:23094070

  12. Effects of Glycine, Water, Ammonia, and Ammonium Bicarbonate on the Oligomerization of Methionine

    NASA Astrophysics Data System (ADS)

    Huang, Rui; Furukawa, Yoshihiro; Otake, Tsubasa; Kakegawa, Takeshi

    2016-09-01

    The abiotic oligomerization of amino acids may have created primordial, protein-like biological catalysts on the early Earth. Previous studies have proposed and evaluated the potential of diagenesis for the amino acid oligomerization, simulating the formation of peptides that include glycine, alanine, and valine, separately. However, whether such conditions can promote the formation of peptides composed of multiple amino acids remains unclear. Furthermore, the chemistry of pore water in sediments should affect the oligomerization and degradation of amino acids and oligomers, but these effects have not been studied extensively. In this study, we investigated the effects of water, ammonia, ammonium bicarbonate, pH, and glycine on the oligomerization and degradation of methionine under high pressure (150 MPa) and high temperature conditions (175 °C) for 96 h. Methionine is more difficult to oligomerize than glycine and methionine dimer was formed in the incubation of dry powder of methionine. Methionine oligomers as long as trimers, as well as methionylglycine and glycylmethionine, were formed under every condition with these additional compounds. Among the compounds tested, the oligomerization reaction rate was accelerated by the presence of water and by an increase in pH. Ammonia also increased the oligomerization rate but consumed methionine by side reactions and resulted in the rapid degradation of methionine and its peptides. Similarly, glycine accelerated the oligomerization rate of methionine and the degradation of methionine, producing water, ammonia, and bicarbonate through its decomposition. With Gly, heterogeneous dimers (methionylglycine and glycylmethionine) were formed in greater amounts than with other additional compounds although smaller amount of these heterogeneous dimers were formed with other additional compounds. These results suggest that accelerated reaction rates induced by water and co-existing reactive compounds promote the oligomerization

  13. Effects of Glycine, Water, Ammonia, and Ammonium Bicarbonate on the Oligomerization of Methionine.

    PubMed

    Huang, Rui; Furukawa, Yoshihiro; Otake, Tsubasa; Kakegawa, Takeshi

    2016-09-23

    The abiotic oligomerization of amino acids may have created primordial, protein-like biological catalysts on the early Earth. Previous studies have proposed and evaluated the potential of diagenesis for the amino acid oligomerization, simulating the formation of peptides that include glycine, alanine, and valine, separately. However, whether such conditions can promote the formation of peptides composed of multiple amino acids remains unclear. Furthermore, the chemistry of pore water in sediments should affect the oligomerization and degradation of amino acids and oligomers, but these effects have not been studied extensively. In this study, we investigated the effects of water, ammonia, ammonium bicarbonate, pH, and glycine on the oligomerization and degradation of methionine under high pressure (150 MPa) and high temperature conditions (175 °C) for 96 h. Methionine is more difficult to oligomerize than glycine and methionine dimer was formed in the incubation of dry powder of methionine. Methionine oligomers as long as trimers, as well as methionylglycine and glycylmethionine, were formed under every condition with these additional compounds. Among the compounds tested, the oligomerization reaction rate was accelerated by the presence of water and by an increase in pH. Ammonia also increased the oligomerization rate but consumed methionine by side reactions and resulted in the rapid degradation of methionine and its peptides. Similarly, glycine accelerated the oligomerization rate of methionine and the degradation of methionine, producing water, ammonia, and bicarbonate through its decomposition. With Gly, heterogeneous dimers (methionylglycine and glycylmethionine) were formed in greater amounts than with other additional compounds although smaller amount of these heterogeneous dimers were formed with other additional compounds. These results suggest that accelerated reaction rates induced by water and co-existing reactive compounds promote the

  14. 63 FR 41290 - Synthetic Methionine From Japan

    Federal Register 2010, 2011, 2012, 2013, 2014

    1998-08-03

    ... COMMISSION Synthetic Methionine From Japan AGENCY: United States International Trade Commission. ACTION: Institution of a five-year review concerning the antidumping duty order on synthetic methionine from Japan... antidumping duty order on synthetic methionine from Japan would be likely to lead to continuation...

  15. Increasing levels of dietary crystalline methionine affect plasma methionine profiles, ammonia excretion, and the expression of genes related to the hepatic intermediary metabolism in rainbow trout (Oncorhynchus mykiss).

    PubMed

    Rolland, Marine; Skov, Peter V; Larsen, Bodil K; Holm, Jørgen; Gómez-Requeni, Pedro; Dalsgaard, Johanne

    2016-08-01

    Strictly carnivorous fish with high requirements for dietary protein, such as rainbow trout (Oncorhynchus mykiss) are interesting models for studying the role of amino acids as key regulators of intermediary metabolism. Methionine is an essential amino acid for rainbow trout, and works as a signalling factor in different metabolic pathways. The study investigated the effect of increasing dietary methionine intake on the intermediary metabolism in the liver of juvenile rainbow trout. For this purpose, five diets were formulated with increasing methionine levels from 0.60 to 1.29% dry matter. The diets were fed in excess for six weeks before three sampling campaigns carried out successively to elucidate (i) the hepatic expression of selected genes involved in lipid, glucose and amino acid metabolism; (ii) the postprandial ammonia excretion; and (iii) the postprandial plasma methionine concentrations. The transcript levels of enzymes involved in lipid metabolism (fatty acid synthase, glucose 6 phosphate dehydrogenase and carnitine palmitoyl transferase 1 a), gluconeogenesis (fructose-1,6-biphosphatase) and amino acid catabolism (alanine amino transferase and glutamate dehydrogenase) were significantly affected by the increase in dietary methionine. Changes in gene expression reflected to some extent the decrease in ammonia excretion (P=0.022) and in the hepatosomatic index (HSI; P<0.001) when dietary methionine increased. Postprandial plasma methionine concentrations correlated positively with the dietary level (P<0.001) at the different sampling points. The study shows that the expression of several genes related to the hepatic intermediary metabolism in rainbow trout responded in a dose-dependent manner to increasing levels of dietary methionine.

  16. Lysine requirement of broiler chicks as affected by protein source and method of statistical evaluation.

    PubMed

    Barbour, G; Latshaw, J D; Bishop, B

    1993-09-01

    1. An experiment was designed to test if the lysine requirement, expressed as g lysine/kg CP, was the same for several protein sources. 2. Groundnut meal, groundnut meal adjusted with indispensable amino acids or sesame meal supplied the dietary CP at 180 g/kg diet. Increments of lysine (1.5 g/kg diet) were added to each of these diets. 3. The gain, food intake and food efficiency responses of broiler chicks were analysed using a quadratic equation and a two-slope method. An estimate of lysine requirements was also obtained from a survey of college students. 4. The different methods produced widely different estimates of lysine requirement. 5. The average lysine requirement was estimated at 50.1 g lysine/kg CP for groundnut meal, 61.7 for adjusted groundnut meal and 54.9 for sesame meal. 6. Reasons for the effect of statistical analysis and protein source on lysine requirement are discussed.

  17. A gene encoding lysine 6-aminotransferase, which forms the beta-lactam precursor alpha-aminoadipic acid, is located in the cluster of cephamycin biosynthetic genes in Nocardia lactamdurans.

    PubMed Central

    Coque, J J; Liras, P; Laiz, L; Martín, J F

    1991-01-01

    A gene (lat) encoding lysine 6-aminotransferase was found upstream of the pcbAB (encoding alpha-aminoadipylcysteinyl-valine synthetase) and pcbC (encoding isopenicillin N synthase) genes in the cluster of early cephamycin biosynthetic genes in Nocardia lactamdurans. The lat gene was separated by a small intergenic region of 64 bp from the 5' end of the pcbAB gene. The lat gene contained an open reading frame of 1,353 nucleotides (71.4% G + C) encoding a protein of 450 amino acids with a deduced molecular mass of 48,811 Da. Expression of DNA fragments carrying the lat gene in Streptomyces lividans led to a high lysine 6-aminotransferase activity which was absent from untransformed S. lividans. The enzyme was partially purified from S. lividans(pULBS8) and showed a molecular mass of 52,800 Da as calculated by Sephadex gel filtration and polyacrylamide gel electrophoresis. DNA sequences which hybridized strongly with the lat gene of N. lactamdurans were found in four cephamycin-producing Streptomyces species but not in four other actinomycetes which are not known to produce beta-lactams, suggesting that the gene is specific for beta-lactam biosynthesis and is not involved in general lysine catabolism. The protein encoded by the lat gene showed similarity to ornithine-5-aminotransferases and N-acetylornithine-5-aminotransferases and contained a pyridoxal phosphate-binding consensus amino acid sequence around Lys-300 of the protein. The evolutionary implications of the lat gene as a true beta-lactam biosynthetic gene are discussed. Images PMID:1917857

  18. Methionine deficiency reduces autophagy and accelerates death in intestinal epithelial cells infected with enterotoxigenic Escherichia coli.

    PubMed

    Tang, Yulong; Tan, Bie; Xiong, Xia; Li, Fengna; Ren, Wenkai; Kong, Xiangfeng; Qiu, Wei; Hardwidge, Philip R; Yin, Yulong

    2015-10-01

    Infections by enterotoxigenic Escherichia coli (ETEC) result in large economic losses to the swine industry worldwide. Dietary supplementation with amino acids has been considered as a potential mechanism to improve host defenses against infection. The goal of this study was to determine whether methionine deprivation alters ETEC interactions with porcine intestinal epithelial cells. IPEC-1 cells were cultured in media with or without L-methionine. Methionine deprivation resulted in enhanced ETEC adhesion and increased both the cytotoxicity and apoptotic responses of IPEC-1 cells infected with ETEC. Methionine deprivation inhibited IPEC-1 cell autophagic responses, suggesting that the increased cytotoxicity of ETEC to methionine-deprived IPEC-1 cells might be due to defects in autophagy.

  19. Metabolic engineering of Escherichia coli for microbial production of L-methionine.

    PubMed

    Huang, Jian-Feng; Liu, Zhi-Qiang; Jin, Li-Qun; Tang, Xiao-Ling; Shen, Zhen-Yang; Yin, Huan-Huan; Zheng, Yu-Guo

    2017-04-01

    L-methionine has attracted a great deal of attention for its nutritional, pharmaceutical, and clinical applications. In this study, Escherichia coli W3110 was engineered via deletion of a negative transcriptional regulator MetJ and over-expression of homoserine O-succinyltransferase MetA together with efflux transporter YjeH, resulting in L-methionine overproduction which is up to 413.16 mg/L. The partial inactivation of the L-methionine import system MetD via disruption of metI made the engineered E. coli ΔmetJ ΔmetI/pTrcA*H more tolerant to high L-ethionine concentration and accumulated L-methionine to a level 43.65% higher than that of E. coli W3110 ΔmetJ/pTrcA*H. Furthermore, deletion of lysA, which blocks the lysine biosynthesis pathway, led to a further 8.5-fold increase in L-methionine titer of E. coli ΔmetJ ΔmetI ΔlysA/pTrcA*H. Finally, addition of Na2 S2 O3 to the media led to an increase of fermentation titer of 11.45%. After optimization, constructed E. coli ΔmetJ ΔmetI ΔlysA/pTrcA*H was able to produce 9.75 g/L L-methionine with productivity of 0.20 g/L/h in a 5 L bioreactor. This novel metabolically tailored strain of E. coli provides an efficient platform for microbial production of L-methionine. Biotechnol. Bioeng. 2017;114: 843-851. © 2016 Wiley Periodicals, Inc.

  20. Methionine kinetics in adult men: effects of dietary betaine on L-(2H3-methyl-1-13C)methionine

    SciTech Connect

    Storch, K.J.; Wagner, D.A.; Young, V.R. )

    1991-08-01

    The effects of a daily 3-g supplement of betaine on kinetic aspects of L-(2H3-methyl-1-13C)methionine (MET) metabolism in healthy young adult men were explored. Four groups of four subjects each were given a control diet, based on an L-amino acid mixture supplying 29.5 and 21.9 mg.kg-1.d-1 of L-methionine and L-cystine for 4 d before the tracer study, conducted on day 5 during the fed state. Two groups received the control diet and two groups received the betaine supplement. Tracer was given intravenously (iv) or orally. The transmethylation rate of MET (TM), homocysteine remethylation (RM), and oxidation of methionine were estimated from plasma methionine labeling and 13C enrichment of expired air. RM tended to increase (P = 0.14) but the TM and methionine oxidation were significantly (P less than 0.05) higher after betaine supplementation when estimated with the oral tracer. No differences were detected with the intravenous tracer. Methionine concentration in plasma obtained from blood taken from subjects in the fed state was higher (P less than 0.01) with betaine supplementation. These results suggest that excess methyl-group intake may increase the dietary requirement for methionine.

  1. Food safety and amino acid balance in processed cassava "Cossettes".

    PubMed

    Diasolua Ngudi, Delphin; Kuo, Yu Haey; Lambein, Fernand

    2002-05-08

    Processed cassava (Manihot esculenta Crantz) roots provide more than 60% of the daily energy intake for the population of the Democratic Republic of Congo. Insufficiently processed cassava roots in a diet deficient in sulfur amino acid have been reported to cause the irreversible paralytic disease konzo, afflicting thousands of women and children in the remote rural areas of Bandundu Province. "Cossettes" (processed cassava roots) purchased in several markets of Kinshasa were analyzed for their content of cyanogens, free amino acids, and total protein amino acids. Residual cyanogen levels were below the safe limit recommended by the codex FAO/WHO for cassava flour (10 mg kg(-1)). The amino acid score was evaluated. Lysine and leucine were the limiting amino acids. Methionine content was very low and contributed about 13% of the total sulfur amino acids. Dietary requirements for sulfur amino acids need to be adjusted for the loss caused by cyanogen detoxification.

  2. Recycling of 5'-methylthioadenosine-ribose carbon atoms into methionine in tomato tissue in relation to ethylene production.

    PubMed

    Wang, S Y; Adams, D O; Lieberman, M

    1982-07-01

    The ribose moiety of 5'-methylthioadenosine (MTA) is metabolized to form the four-carbon unit (2-aminobutyrate) of methionine in tomato tissue (Lycopersicon esculentum Mill., cv. Pik Red). When [U-(14)C-adenosine] MTA was administered to tomato tissue slices, label was recovered in 5-methylthioribose (MTR), methionine, 1-aminocyclopropane-1-carboxylic acid (ACC), C(2)H(4) and other unidentified compounds. However, when [U-(14)C-ribose]MTR was administered, radioactivities were recovered in methionine, ACC and C(2)H(4), but not MTA. This suggests that C(2)H(4) formed in tomato pericarp tissue may be derived from the ribose portion of MTA via MTR, methionine and ACC. The conversion of MTR to methionine is not inhibited by aminoethoxyvinylglycine (AVG), but is O(2) dependent. These data present a new salvage pathway for methionine biosynthesis which may be important in relation to polyamine and ethylene biosynthesis in tomato tissue.

  3. Conformation of Lysine Vasopressin: A Comparison with Oxytocin

    PubMed Central

    Walter, Roderich; Glickson, J. D.; Schwartz, I. L.; Havran, R. T.; Meienhofer, Johannes; Urry, D. W.

    1972-01-01

    Starting with assignments of proton nuclear magnetic resonance previously made for oxytocin in deuterated dimethylsulfoxide at 220 MHz, we have assigned resonances for the mammalian antidiuretic hormone, lysine vasopressin. The results demonstrate that spectral assignments of neurohypophyseal hormones and their congeners can, within certain limits, be derived from each other. Comparison of the spectra of lysine vasopressin and oxytocin suggests that the gross backbone conformations of their 20-membered ring components are for the most part similar in deuterated dimethylsulfoxide, whereas the C-terminal acyclic amino-acid sequence of lysine vasopressin is more flexible than that of oxytocin. PMID:4505670

  4. Mutants of Saccharomycopsis lipolytica defective in lysine catabolism.

    PubMed Central

    Gaillardin, C; Fournier, P; Sylvestre, G; Heslot, H

    1976-01-01

    Wild-type strains of Saccharomycopsis lipolytica are able to use lysine as a carbon or a nitrogen source, but not as a unique source for both. Mutants were selected that could not use lysine either as a nitrogen or as a carbon source. Some of them, however, utilized N-6-acetyllysine or 5-aminovaleric acid. Many of the mutants appeared to be blocked in both utilizations, suggesting a unique pathway for lysine degradation (either as a carbon or as a nitrogen source). Genetic characterization of these mutants was achieved by complementation and recombination tests. PMID:1245461

  5. Methionine sulfoximine intensifies cancer anorexia.

    PubMed

    Chance, W T; Zhang, F S; Fischer, J E

    1991-05-01

    Consistent anorexia was first observed 33 days after inoculating Fischer 344 rats with methylcholanthrene-induced sarcoma. Daily treatment of a similar group of rats with the glutamine synthetase inhibitor, methionine sulfoximine, elicited significant reductions of feeding by day 29 at a dose that had no effect on nontumor-bearing rats. Blood concentrations of ammonia were elevated in both groups of tumor-bearing rats and brain ammonia level was increased in the methionine sulfoximine-treated tumor-bearing rats. Forebrain concentrations of tyrosine, tryptophan, DOPAC and 5-HIAA were elevated in both groups of tumor-bearing rats. Since ammonia is detoxified through the glutamine synthetase reaction, these results suggest that blood and brain ammonia concentrations are more important than the neurochemical consequences of ammonia detoxification for the etiology of cancer anorexia.

  6. The contents of the neuro-excitatory amino acid β-ODAP (β-N-oxalyl-l-α,β-diaminopropionic acid), and other free and protein amino acids in the seeds of different genotypes of grass pea (Lathyrus sativus L.).

    PubMed

    Fikre, Asnake; Korbu, Lijalem; Kuo, Yu-Haey; Lambein, Fernand

    2008-09-15

    The free and protein amino acids of nine different genotypes of grass pea (Lathyrus sativus L.) seeds were analysed by HPLC with pre-column PITC (phenyl isothiocyanate) derivatisation. Among the free amino acids, homoarginine was quantitatively the most important (up to 0.8% seed weight) and stable while the neuro-excitatory amino acid β-ODAP (β-N-oxalyl-l-α,β-diaminopropionic acid) showed highest variation (0.02-0.54%) in the nine genotypes examined. Among protein amino acids, glutamic acid was quantitatively most significant, followed by aspartic acid, arginine, leucine, lysine and proline. The sulphur amino acid, methionine, showed the lowest concentration in all the L. sativus genotypes, and also in lentil (Lens culinaris) and in soybean (Glycine max) seeds analysed at the same time.

  7. Modified bean seed protein phaseolin did not accumulate stably in transgenic tobacco seeds after methionine enhancement mutations

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The major seed storage protein phaseolin of common bean (Phaseolus vulgaris L.) is deficient in methionine, an essential amino acid for human and animal health. To improve the nutritional quality of common bean, we designed methionine enhancement of phaseolin based on the three dimensional structure...

  8. The reaction of methionine with hydroxyl radical: reactive intermediates and methanethiol production.

    PubMed

    Spasojević, Ivan; Bogdanović Pristov, Jelena; Vujisić, Ljubodrag; Spasić, Mihajlo

    2012-06-01

    The mechanisms of reaction of methionine with hydroxyl radical are not fully understood. Here, we unequivocally show using electron paramagnetic resonance spin-trapping spectroscopy and GC-FID and GC-MS, the presence of specific carbon-, nitrogen- and sulfur-centered radicals as intermediates of this reaction, as well as the liberation of methanethiol as a gaseous end product. Taking into account the many roles that methionine has in eco- and biosystems, our results may elucidate redox chemistry of this amino acid and processes that methionine is involved in.

  9. Na-Stimulated Transport of l-Methionine in Brevibacterium linens CNRZ 918.

    PubMed

    Ferchichi, M; Hemme, D; Nardi, M

    1987-09-01

    The transport of l-methionine by the gram-positive species Brevibacterium linens CNRZ 918 is described. The one transport system (K(m) = 55 muM) found is constitutive for l-methionine, stereospecific, and pH and temperature dependent. Entry of l-methionine into cells is controlled by the internal methionine pool. Competition studies indicate that l-methionine and alpha-aminobutyric acid share a common carrier for their transport. Neither methionine derivatives substituted on the amino or carboxyl groups nor d-methionine was an inhibitor, whereas powerful inhibition was shown by l-cysteine, s-methyl-l-cysteine, dl-selenomethionine and dl-homocysteine. Sodium plays important and varied roles in l-methionine transport by B. linens CNRZ 918: (i) it stimulates transport without affecting the K(m), (ii) it increases the specific activity (on a biomass basis) of the l-methionine transport system when present with methionine in the medium, suggesting a coinduction mechanism. l-Methionine transport requires an exogenous energy source, which may be succinic, lactic, acetic, or pyruvic acid but not glucose or sucrose. The fact that l-methionine transport was stimulated by potassium arsenate and to a lesser extent by potassium fluoride suggests that high-energy phosphorylated intermediates are not involved in the process. Monensin eliminates stimulation by sodium. Gramicidin and carbonyl cyanide-m-chlorophenylhydrazone act in the presence or absence of Na. N-Ethylmaleimide, p-chloromercurobenzoate, valinomycin, sodium azide, and potassium cyanide have no or only a partial inhibitory effect. These results tend to indicate that the proton motive force reinforced by the Na gradient is involved in the mechanism of energy coupling of l-methionine transport by B. linens CNRZ 918. Thus, this transport is partially similar to the well-described systems in gram-negative bacteria, except for the role of sodium, which is very effective in B. linens, a species adapted to the high sodium

  10. Arginine and Lysine Transporters Are Essential for Trypanosoma brucei

    PubMed Central

    Hürlimann, Daniel; Wirdnam, Corina; Haindrich, Alexander C.; Suter Grotemeyer, Marianne; González-Salgado, Amaia; Schmidt, Remo S.; Inbar, Ehud; Mäser, Pascal; Bütikofer, Peter; Zilberstein, Dan; Rentsch, Doris

    2017-01-01

    For Trypanosoma brucei arginine and lysine are essential amino acids and therefore have to be imported from the host. Heterologous expression in Saccharomyces cerevisiae mutants identified cationic amino acid transporters among members of the T. brucei AAAP (amino acid/auxin permease) family. TbAAT5-3 showed high affinity arginine uptake (Km 3.6 ± 0.4 μM) and high selectivity for L-arginine. L-arginine transport was reduced by a 10-times excess of L-arginine, homo-arginine, canavanine or arginine-β-naphthylamide, while lysine was inhibitory only at 100-times excess, and histidine or ornithine did not reduce arginine uptake rates significantly. TbAAT16-1 is a high affinity (Km 4.3 ± 0.5 μM) and highly selective L-lysine transporter and of the compounds tested, only L-lysine and thialysine were competing for L-lysine uptake. TbAAT5-3 and TbAAT16-1 are expressed in both procyclic and bloodstream form T. brucei and cMyc-tagged proteins indicate localization at the plasma membrane. RNAi-mediated down-regulation of TbAAT5 and TbAAT16 in bloodstream form trypanosomes resulted in growth arrest, demonstrating that TbAAT5-mediated arginine and TbAAT16-mediated lysine transport are essential for T. brucei. Growth of induced RNAi lines could partially be rescued by supplementing a surplus of arginine or lysine, respectively, while addition of both amino acids was less efficient. Single and double RNAi lines indicate that additional low affinity uptake systems for arginine and lysine are present in T. brucei. PMID:28045943

  11. Effects of Graded Levels of Chromium Methionine on Performance, Carcass Traits, Meat Quality, Fatty Acid Profiles of Fat, Tissue Chromium Concentrations, and Antioxidant Status in Growing-Finishing Pigs.

    PubMed

    Tian, Yao-Yao; Gong, Li-Min; Xue, Jian-Xiang; Cao, Jun; Zhang, Li-Ying

    2015-11-01

    A 97-day feeding trial was conducted to investigate the effects of dietary chromium methionine (CrMet) on performance, carcass traits, meat quality, fatty acid profiles of fat, tissue chromium concentrations, and antioxidant status in growing-finishing pigs. A total of 180 crossbred pigs with a mean initial body weight (BW) 30.18 ± 0.28 kg were allotted to 5 treatments with 6 replicates per treatment and 6 pigs per pen in a randomized complete block design based on BW and sex. Treatments were added with 0 (control), 100, 200, 400, and 800 μg/kg chromium as CrMet. Blood samples were obtained from the anterior vena cava on days 97. Carcass characteristics, pork quality, and tissue chromium concentration data were collected from one pig per pen. The results indicated that supplemental CrMet did not significantly affect growth performance, carcass traits, or meat amino acid profiles. Chromium at 100, 400, and 800 μg/kg decreased drip loss but increased shear force (P < 0.05). Pigs fed 100 or 400 μg/kg had a higher 24-h pH than the control (P < 0.05). While meat color, muscle moisture, crude protein, or crude fat were not affected by CrMet. Supplemental 800 μg/kg chromium reduced C18:0 levels in belly fat (P < 0.05), and chromium supplementation increased cis-9, trans 11-conjugated linoleic acid levels linearly (P < 0.05). Dietary CrMet supplementation increased serum, kidney, and muscle chromium contents (P < 0.05) but did not affect liver chromium contents. Besides, tissue chromium concentrations were increased linearly with increased chromium dosage (P < 0.05). Chromium at 400 μg/kg increased serum glutathione peroxidase activities (P < 0.05), and chromium at 800 μg/kg decreased serum total antioxidant capacity levels (P < 0.05). Nevertheless, liver and kidney antioxidant status were not significantly affected by CrMet. These results indicated that dietary supplementation CrMet did not significantly influence growth

  12. (11)C-Methionine uptake in secondary brain epilepsy.

    PubMed

    Lopci, E; Bello, L; Chiti, A

    2014-01-01

    Carbon-11 methionine ((11)C-Methionine) is a radio-labeled amino acid currently utilized in Positron Emission Tomography (PET) for imaging primary and metastatic brain tumors. Its clinical use relies mostly on oncologic applications, but the tracer has the potential to investigate other non-malignant conditions. So far, very limited evidence concerns the use of (11)C-Methionine in patients suffering from seizure; however, the tracer can find a proper utilization in this setting especially as a diagnostic complement to (18)F-Fluorodeoxyglucose ((18)F-FDG). Herein we report the case of a 57-year-old patient presenting with epileptic crises secondary to a brain metastasis from bladder carcinoma, who was investigated in our institution with (11)C-Methionine PET. The scan documented the disease recurrence in the left parietal lobe associated with a diffused tracer uptake in the surrounding cerebral circumvolutions, derived from the comitial status. After surgical removal of the metastatic lesion, the patient experienced a complete recovery of symptoms and no further onset of secondary seizure.

  13. Technical note: Methionine, a precursor of methane in living plants

    NASA Astrophysics Data System (ADS)

    Lenhart, K.; Althoff, F.; Greule, M.; Keppler, F.

    2014-11-01

    When terrestrial plants were identified as producers of the greenhouse gas methane, much discussion and debate ensued, not only about their contribution to the global methane budget, but also with regard to the validity of the observation itself. Although the phenomenon has now become more accepted for both living and dead plants, the mechanism of methane formation in living plants remains to be elucidated and its precursor compounds identified. We made use of stable isotope techniques to verify in vivo formation of methane and, in order to identify the carbon precursor, 13C-positionally labelled organic compounds were employed. Here we show that the amino acid L-methionine acts as a methane precursor in living plants. Employing 13C-labelled methionine clearly identified the sulphur-bound methyl group of methionine as a carbon precursor of methane released from lavender (Lavandula angustifolia). Furthermore, when lavender plants were stressed physically, methane release rates and the stable carbon isotope values of the emitted methane greatly increased. Our results provide additional support that plants possess a mechanism for methane production and suggest that methionine might play an important role in the formation of methane in living plants, particularly under stress conditions.

  14. Technical Note: Methionine, a precursor of methane in living plants

    NASA Astrophysics Data System (ADS)

    Lenhart, K.; Althoff, F.; Greule, M.; Keppler, F.

    2015-03-01

    When terrestrial plants were identified as producers of the greenhouse gas methane, much discussion and debate ensued not only about their contribution to the global methane budget but also with regard to the validity of the observation itself. Although the phenomenon has now become more accepted for both living and dead plants, the mechanism of methane formation in living plants remains to be elucidated and its precursor compounds to be identified. We made use of stable isotope techniques to verify the in vivo formation of methane, and, in order to identify the carbon precursor, 13C positionally labeled organic compounds were employed. Here we show that the amino acid L-methionine acts as a methane precursor in living plants. Employing 13C-labeled methionine clearly identified the sulfur-bound methyl group of methionine as a carbon precursor of methane released from lavender (Lavandula angustifolia). Furthermore, when lavender plants were stressed physically, methane release rates and the stable carbon isotope values of the emitted methane greatly increased. Our results provide additional support that plants possess a mechanism for methane production and suggest that methionine might play an important role in the formation of methane in living plants, particularly under stress conditions.

  15. Metabolism of methionine and biosynthesis of caffeine in the tea plant (Camellia sinensis L.).

    PubMed Central

    Suzuki, T; Takahashi, E

    1976-01-01

    1. Caffeine biosynthesis was studied by following the incorporation of 14C into the products of L-[Me-14C]methionine metabolism in tea shoot tips. 2. After administration of a 'pulse' of L-[Me-14C]methionine, almost all of the L-[Me-14C]methionine supplied disappeared within 1 h, and 14C-labelled caffeine synthesis increased throughout the experimental periods, whereas the radioactivities of an unknown compound and theobromine were highest at 3 h after the uptake of L-[Me-14C]methionine, followed by a steady decrease. There was also slight incorporation of the label into 7-methylxanthine, serine, glutamate and aspartate, disappearing by 36 h after the absorption of L-[Me-14C]methionine. 3. The radioactivities of nucleic acids derived from L-[Me-14C]methionine increased rapidly during the first 12 h incubation period and then decreased steadily. Sedimentation analysis of nucleic acids by sucrose-gradient centrifugation showed that methylation of nucleic acids in tea shoot tips occurred mainly in the tRNA fraction. The main product among the methylated bases in tea shoot tips was identified as 1-methyladenine. 4. The results indicated that the purine ring in caffeine is derived from the purine nucleotides in the nucleotide pool rather than in nucleic acids. A metabolic scheme to show the production of caffeine and related methylxanthines from the nucleotides in tea plants is discussed. PMID:1008848

  16. Simultaneous determination of plasma total homocysteine and methionine by liquid chromatography-tandem mass spectrometry.

    PubMed

    Jiang, Yi; Mistretta, Brandon; Elsea, Sarah; Sun, Qin

    2017-01-01

    The sulfur-containing amino acid homocysteine is a cardiac risk factor and a biomarker for several inborn errors of metabolism in methionine synthesis. A simple LC-MS/MS method was developed and validated for determination of homocysteine and methionine in human plasma. Rapid separation was achieved using a reverse phase liquid chromatography. Mass spectrometry identification was performed in positive electrospray ionization mode for homocysteine and methionine. Accuracy, precision, linearity, recovery and sample stability were evaluated in the method validation. The test is applied in diagnosis of homocystinuria and monitoring total homocysteine levels. Moreover, simultaneous measurement of methionine helps in the differentiation of homocystinuria and some cobalamin disorders (such as cblC and cblD defects) without additional amino acid testing. Lastly, this assay is sensitive to detect reduced total homocysteine levels that are possibly seen in sulfocysteinuria and molybdenum cofactor deficiencies.

  17. Effects of methyl-deficient diets on methionine and homocysteine metabolism in the pregnant rat.

    PubMed

    Wilson, Fiona A; Holtrop, Grietje; Calder, A Graham; Anderson, Susan E; Lobley, Gerald E; Rees, William D

    2012-06-15

    Although the importance of methyl metabolism in fetal development is well recognized, there is limited information on the dynamics of methionine flow through maternal and fetal tissues and on how this is related to circulating total homocysteine concentrations. Rates of homocysteine remethylation in maternal and fetal tissues on days 11, 19, and 21 of gestation were measured in pregnant rats fed diets with limiting or surplus amounts of folic acid and choline at two levels of methionine and then infused with L-[1-(13)C,(2)H(3)-methyl]methionine. The rate of homocysteine remethylation was highest in maternal liver and declined as gestation progressed. Diets deficient in folic acid and choline reduced the production of methionine from homocysteine in maternal liver only in the animals fed a methionine-limited diet. Throughout gestation, the pancreas exported homocysteine for methylation within other tissues. Little or no methionine cycle activity was detected in the placenta at days 19 and 21 of gestation, but, during this period, fetal tissues, especially the liver, synthesized methionine from homocysteine. Greater enrichment of homocysteine in maternal plasma than placenta, even in animals fed the most-deficient diets, shows that the placenta did not contribute homocysteine to maternal plasma. Methionine synthesis from homocysteine in fetal tissues was maintained or increased when the dams were fed folate- and choline-deficient methionine-restricted diets. This study shows that methyl-deficient diets decrease the remethylation of homocysteine within maternal tissues but that these rates are protected to some extent within fetal tissues.

  18. Solubility Behavior of Cyanophycin Depending on Lysine Content

    PubMed Central

    Wiefel, Lars

    2014-01-01

    Study of the synthesis of cyanophycin (CGP) in recombinant organisms focused for a long time mostly on the insoluble form of CGP, due to its easy purification and its putative use as a precursor for biodegradable chemicals. Recently, another form of CGP, which, in contrast to the insoluble form, was soluble at neutral pH, became interesting due to its high lysine content, which was also assumed to be the reason for the solubility of the polymer. In this study, we demonstrate that lysine incorporated into insoluble CGP affected the solubility of the polymer in relation to its lysine content. Insoluble CGP can be separated along a temperature gradient of 90°C to 30°C, where CGP showed an increasing lysine content corresponding to a decreasing temperature needed for solubilization. CGP with less than 3 to 4 mol% lysine did not become soluble even at 90°C, while CGP with 31 mol% lysine was soluble at 30°C. In lysine fractions at higher than 31 mol%, CGP was soluble. The temperature separation will be suitable for improving the downstream processing of CGP synthesized in large-scale fermentations, including faster and more efficient purification of CGP, as well as enrichment and separation of dipeptides and CGP with specific amino acid compositions. PMID:24271185

  19. A novel potentiometric biosensor for determination of L-lysine in commercial pharmaceutical L-lysine tablet and capsule.

    PubMed

    Yarar, Saniye; Karakuş, Emine

    2016-01-01

    The construction of an L-lysine biosensor on ammonium-selective poly(vinylchloride) (PVC) membrane electrode is described in this study. The construction procedure occurs in two stages: (I) the preparation of ammonium-selective poly(vinylchloride) (PVC) membrane electrode and (II) the chemical immobilization of lysine oxidase on this ammonium-selective electrode by using glutaraldehyde. The ammonium ions produced after enzymatic reaction were determined potentiometrically. The sensitivity of the lysine biosensor against ammonium ions and lysine were studied. The response time, linear working range, reproducibility and life time of the biosensor were also determined. The interfering effect of other amino acids on the biosensor performance was also studied and potentiometric selectivity coefficients were calculated. Although the biosensor responded mainly against tyrosine, a lot of amino acids and ascorbic acid that can be present in some real samples did not show any important interference. Additionally, lysine assay in commercial pharmaceutical lysine tablets and capsules was also successfully carried out. The results were in good agreement with previously reported values.

  20. Free amino acid profiling in the giant puffball mushroom (Calvatia gigantea) using UPLC-MS/MS.

    PubMed

    Kıvrak, İbrahim; Kıvrak, Şeyda; Harmandar, Mansur

    2014-09-01

    Wild edible and medicinal mushroom, Calvatia gigantea, was quantitatively analyzed for the determination of its free amino acids using ultra performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS). The concentrations of total free amino acids, essential and non-essential amino acids were 199.65 mg/100 g, 113.69 mg/100 g, and 85.96 mg/100 g in C. gigantea, respectively. This study showed that C. gigantea, so called a giant puffball mushroom, has free amino acids content. The essential amino acids: tryptophan, isoleucine, valine, phenylalanine, leucine, threonine, lysine, histidine, methionine, and the non-essential amino acids: tyrosine, 4-hyrdroxy proline, arginine, proline, glycine, serine, alanine, glutamine, glutamic acid, aspargine, aspartic acid were detected.

  1. Distinguishing the cyanobacterial neurotoxin β-N-methylamino-L-alanine (BMAA) from other diamino acids.

    PubMed

    Banack, S A; Metcalf, J S; Spáčil, Z; Downing, T G; Downing, S; Long, A; Nunn, P B; Cox, P A

    2011-04-01

    β-N-methylamino-L-alanine (BMAA) is produced by diverse taxa of cyanobacteria, and has been detected by many investigators who have searched for it in cyanobacterial blooms, cultures and collections. Although BMAA is distinguishable from proteinogenic amino acids and its isomer 2,4-DAB using standard chromatographic and mass spectroscopy techniques routinely used for the analysis of amino acids, we studied whether BMAA could be reliably distinguished from other diamino acids, particularly 2,6-diaminopimelic acid which has been isolated from the cell walls of many bacterial species. We used HPLC-FD, UHPLC-UV, UHPLC-MS, and triple quadrupole tandem mass spectrometry (UHPLC-MS/MS) to differentiate BMAA from the diamino acids 2,6-diaminopimelic acid, N-2(amino)ethylglycine, lysine, ornithine, 2,4-diaminosuccinic acid, homocystine, cystine, tryptophan, as well as other amino acids including asparagine, glutamine, and methionine methylsulfonium.

  2. Growth hormone signaling is necessary for lifespan extension by dietary methionine.

    PubMed

    Brown-Borg, Holly M; Rakoczy, Sharlene G; Wonderlich, Joseph A; Rojanathammanee, Lalida; Kopchick, John J; Armstrong, Vanessa; Raasakka, Debbie

    2014-12-01

    Growth hormone significantly impacts lifespan in mammals. Mouse longevity is extended when growth hormone (GH) signaling is interrupted but markedly shortened with high-plasma hormone levels. Methionine metabolism is enhanced in growth hormone deficiency, for example, in the Ames dwarf, but suppressed in GH transgenic mice. Methionine intake affects also lifespan, and thus, GH mutant mice and respective wild-type littermates were fed 0.16%, 0.43%, or 1.3% methionine to evaluate the interaction between hormone status and methionine. All wild-type and GH transgenic mice lived longer when fed 0.16% methionine but not when fed higher levels. In contrast, animals without growth hormone signaling due to hormone deficiency or resistance did not respond to altered levels of methionine in terms of lifespan, body weight, or food consumption. Taken together, our results suggest that the presence of growth hormone is necessary to sense dietary methionine changes, thus strongly linking growth and lifespan to amino acid availability.

  3. Topological dispositions of lysine. alpha. 380 and lysine. gamma. 486 in the acetylcholine receptor from Torpedo californica

    SciTech Connect

    Dwyer, B.P. )

    1991-04-23

    The locations have been determined, with respect to the plasma membrane, of lysine {alpha}380 and lysine {gamma}486 in the {alpha} subunit and the {gamma} subunit, respectively, of the nicotinic acetylcholine receptor from Torpedo californica. Immunoadsorbents were constructed that recognize the carboxy terminus of the peptide GVKYIAE released by proteolytic digestion from positions 378-384 in the amino acid sequence of the {alpha} subunit of the acetylcholine receptor and the carboxy terminus of the peptide KYVP released by proteolytic digestion from positions 486-489 in the amino acid sequence of the {gamma} subunit. They were used to isolate these peptides from proteolytic digests of polypeptides from the acetylcholine receptor. Sealed vesicles containing the native acetylcholine receptor were labeled with pyridoxal phosphate and sodium ({sup 3}H)-borohydride. The effect of saponin on the incorporation of pyridoxamine phosphate into lysine {alpha}380 and lysine {gamma}486 from the acetylcholine receptor in these vesicles was assessed with the immunoadsorbents. The conclusions that follow from these results are that lysine {alpha}380 is on the inside surface of a vesicle and lysine {gamma}486 is on the outside surface. Because a majority (85%) of the total binding sites for {alpha}-bungarotoxin bind the toxin in the absence of saponin, the majority of the vesicles are right side out with the inside of the vesicle corresponding to the cytoplasmic surface and the outside of the vesicle corresponding to the extracytoplasmic, synaptic surface. Because lysine {alpha}380 and lysine {gamma}486 lie on opposite sides of the membrane, a membrane-spanning segment must be located between the two positions occupied by these two amino acids in the common sequence of a polypeptide of the acetylcholine receptor.

  4. Methionine uptake in Corynebacterium glutamicum by MetQNI and by MetPS, a novel methionine and alanine importer of the NSS neurotransmitter transporter family.

    PubMed

    Trötschel, Christian; Follmann, Martin; Nettekoven, Jeannine A; Mohrbach, Tobias; Forrest, Lucy R; Burkovski, Andreas; Marin, Kay; Krämer, Reinhard

    2008-12-02

    The soil bacterium Corynebacterium glutamicum is a model organism in amino acid biotechnology. Here we present the identification of two different L-methionine uptake systems including the first characterization of a bacterial secondary methionine carrier. The primary carrier MetQNI is a high affinity ABC-type transporter specific for l-methionine. Its expression is under the control of the transcription factor McbR, the global regulator of sulfur metabolism in C. glutamicum. Besides MetQNI, a novel secondary methionine uptake system of the NSS (neurotransmitter:sodium symporter) family was identified and named MetP. The MetP system is characterized by a lower affinity for methionine and uses Na(+) ions for energetic coupling. It is also the main alanine transporter in C. glutamicum and is expressed constitutively. These observations are consistent with models of methionine, alanine, and leucine bound to MetP, derived from the X-ray crystal structure of the LeuT transporter from Aquifex aeolicus. Complementation studies show that MetP consists of two components, a large subunit with 12 predicted transmembrane segments and, surprisingly, an additional subunit with one predicted transmembrane segment only. Thus, this new member of the NSS transporter family adds a novel feature to this class of carriers, namely, the functional dependence on an additional small subunit.

  5. Efficiency of methionine retention in ducks.

    PubMed

    Adeola, Olayiwola

    2007-03-01

    The accretion of methionine and protein as a function of methionine intake was assessed in growing ducks between 22 and 42 d post-hatching. Four graded doses of dl-methionine at 0, 0 x 5, 1 x 0 or 1 x 5 g/kg diet were added to a methionine-limiting basal diet and fed to four replicate groups of four ducks each. The growth and efficiency of food use for growth increased linearly (P<0 x 05) as a function of methionine intake. The accretion of body protein increased (P<0 x 001) from 87 x 5 to 182 x 2 g, and that of methionine from 1616 to 3125 mg, over the 21 d period as dietary methionine increased. The accretion rate of methionine in the body (y, mg/d) as a function of methionine intake (x, mg/d) of ducks fed diets containing supplemental methionine at 0, 0 x 5, 1 x 0 or 1 x 5 g/kg diet from day 22 to day 42 post-hatching gave the regression equation: y=-148 x 86 (se 32 x 558)+0 x 312 (se 0 x 0384)X, r2=0 x 8253. For protein accretion rate in the body (y, mg/d) as a function of methionine intake (x, mg/d), the regression equation was: y= -9782 (se 2204)+19 x 505 (se 2 x 5994)x, r2=0 x 8009. There was a linear relationship between methionine (y, mg/d) and protein (x, mg/d) accretion in ducks that was described by the equation y=12 x 757 (se 7 x 4019)+0 x 01 525 (se 0 x 00 107)x, r2=0 x 9355. The results of these studies suggest a constant utilisation of methionine over the range 2 x 4-3 x 9 g digestible methionine/kg diet, with an efficiency of 31 %. Furthermore, the results suggest a quantitative relationship of 15 mg methionine for every gram of protein accretion.

  6. Biofortification of rice with lysine using endogenous histones.

    PubMed

    Wong, H W; Liu, Q; Sun, S S M

    2015-02-01

    Rice is the most consumed cereal grain in the world, but deficient in the essential amino acid lysine. Therefore, people in developing countries with limited food diversity who rely on rice as their major food source may suffer from malnutrition. Biofortification of stable crops by genetic engineering provides a fast and sustainable method to solve this problem. In this study, two endogenous rice lysine-rich histone proteins, RLRH1 and RLRH2, were over-expressed in rice seeds to achieve lysine biofortification. Their protein sequences passed an allergic sequence-based homology test. Their accumulations in rice seeds were raised to a moderate level by the use of a modified rice glutelin 1 promoter with lowered expression strength to avoid the occurrence of physiological abnormalities like unfolded protein response. The expressed proteins were further targeted to protein storage vacuoles for stable storage using a glutelin 1 signal peptide. The lysine content in the transgenic rice seeds was enhanced by up to 35 %, while other essential amino acids remained balanced, meeting the nutritional standards of the World Health Organization. No obvious unfolded protein response was detected. Different degrees of chalkiness, however, were detected in the transgenic seeds, and were positively correlated with both the levels of accumulated protein and lysine enhancement. This study offered a solution to the lysine deficiency in rice, while at the same time addressing concerns about food safety and physiological abnormalities in biofortified crops.

  7. Formation of methionine sulfoxide during glycoxidation and lipoxidation of ribonuclease A.

    PubMed

    Brock, Jonathan W C; Ames, Jennifer M; Thorpe, Suzanne R; Baynes, John W

    2007-01-15

    Chemical modification of proteins by reactive oxygen species affects protein structure, function and turnover during aging and chronic disease. Some of this damage is direct, for example by oxidation of amino acids in protein by peroxide or other reactive oxygen species, but autoxidation of ambient carbohydrates and lipids amplifies both the oxidative and chemical damage to protein and leads to formation of advanced glycoxidation and lipoxidation end-products (AGE/ALEs). In previous work, we have observed the oxidation of methionine during glycoxidation and lipoxidation reactions, and in the present work we set out to determine if methionine sulfoxide (MetSO) in protein was a more sensitive indicator of glycoxidative and lipoxidative damage than AGE/ALEs. We also investigated the sites of methionine oxidation in a model protein, ribonuclease A (RNase), in order to determine whether analysis of the site specificity of methionine oxidation in proteins could be used to indicate the source of the oxidative damage, i.e. carbohydrate or lipid. We describe here the development of an LC/MS/MS for quantification of methionine oxidation at specific sites in RNase during glycoxidation or lipoxidation by glucose or arachidonate, respectively. Glycoxidized and lipoxidized RNase were analyzed by tryptic digestion, followed by reversed phase HPLC and mass spectrometric analysis to quantify methionine and methionine sulfoxide containing peptides. We observed that: (1) compared to AGE/ALEs, methionine sulfoxide was a more sensitive biomarker of glycoxidative or lipoxidative damage to proteins; (2) regardless of oxidizable substrate, the relative rate of oxidation of methionine residues in RNase was Met29>Met30>Met13, with Met79 being resistant to oxidation; and (3) arachidonate produced a significantly greater yield of MetSO, compared to glucose. The methods developed here should be useful for assessing a protein's overall exposure to oxidative stress from a variety of sources in

  8. Metabolic characteristics and importance of the universal methionine salvage pathway recycling methionine from 5'-methylthioadenosine.

    PubMed

    Albers, Eva

    2009-12-01

    The methionine salvage pathway, also called the 5'-methylthioadenosine (MTA) cycle, recycles the sulfur of MTA, which is a by-product in the biosyntheses of polyamine and the plant hormone ethylene. MTA is first converted to 5'-methylthioribose-1-phosphate either by MTA phosphorylase or the combined action of MTA nucleosidase and 5'-methylthioribose kinase. Subsequently, five additional enzymatic steps, catalyzed by four or five proteins, will form 4-methylthio-2-oxobutyrate, the deaminated form of methionine. The final transamination is achieved by transaminases active in the amino acid biosynthesis. This pathway is present with some variations in all types of organisms and seems to be designed for a quick removal of MTA achieved by high affinities of the first enzymes. During evolution some enzymes have attained additional functions, like a proposed role in nuclear mRNA processing by the aci-reductone dioxygenase. For others the function seems to be lost due to conditions in specific ecological niches, such as, presence of sulfur and/or absence of oxygen resulting in that, for example, Escherichia coli is lacking a functional pathway. The pathway is regulated as response to sulfur availability and take part in the regulation of polyamine synthesis. Some of the enzymes in the pathway show separate specificities in different organisms and some others are unique for groups of bacteria and parasites. Thus, promising targets for antimicrobial agents have been identified. Other medical topics to which this pathway has connections are cancer, apoptosis, and inflammatory response.

  9. Metabolism of excess methionine in the liver of intact rat: an in vivo /sup 2/H NMR study

    SciTech Connect

    London, R.E.; Gabel, S.A.; Funk, A.

    1987-11-03

    L-Methionine is the most toxic amino acid if supplied in excess, and the metabolic basis for this toxicity has been extensively studied, with varying conclusions. It is demonstrated here that in vivo /sup 2/H NMR spectroscopy provides a useful approach to the study of the hepatic metabolism of methionine in the anesthetized rat. Resonances corresponding to administered L-(methyl-/sup 2/H/sub 3/)methionine, and to the transmethylation product sarcosine, are observed during the first 10-min period after an intravenous injection of the labeled methionine, and the time dependence has been followed for a period of 5 h. A third resonance, assigned to the N-trimethyl groups of carnitine, phosphorylcholine, and other metabolites, becomes observable several hours after administration of the deuteriated methionine. In addition, there is a small increase in the intensity of the HDO resonance over the period of the study, which is interpreted to reflect the ultimate oxidation of the labeled sarcosine methyl group via mitochondrial sarcosine dehydrogenase. Additional small /sup 2/H resonances assigned to N/sup 1/-methylhistidine and creatine could be observed in perchloric acid extracts of the livers of rats treated with the deuteriated methionine. Inhibition of the flux through the transmethylation pathway is observed in the rat pretreated with the S-ethyl analogue of methionine, ethionine. These data provide strong support for the importance of glycine transmethylation in the catabolism of excess methionine.

  10. Effects of amino acids on the physiochemical properties of potato starch.

    PubMed

    Cui, Min; Fang, Ling; Zhou, Hongxian; Yang, Hong

    2014-05-15

    The objective of this study was to evaluate effects of different amino acid additives (phenylalanine (Phe), methionine (Met), lysine (Lys), arginine (Arg), aspartic acid (Asp) and glutamic acid (Glu)) on the physicochemical properties of potato starch gels. Charge-carrying amino acids (Lys, Arg, Asp and Glu) significantly decreased the swelling power, solubility, light transmittance, L(∗) value and gel strength of potato starch, but increased syneresis during freeze-thaw treatment, while neutral amino acids (Phe and Met) did not cause modifications in starch gels. During heating, potato starch with fortified charge-carrying amino acids showed a lower peak G' (storage modulus), when compared with Phe and Met. Results showed that charge-carrying amino acids could modify physicochemical properties and improve the nutritional values of starch-based products.

  11. Methionine and choline regulate the metabolic phenotype of a ketogenic diet.

    PubMed

    Pissios, Pavlos; Hong, Shangyu; Kennedy, Adam Richard; Prasad, Deepthi; Liu, Fen-Fen; Maratos-Flier, Eleftheria

    2013-01-01

    Low-carbohydrate ketogenic diets are commonly used as weight loss alternatives to low-fat diets, however the physiological and molecular adaptations to these diets are not completely understood. It is assumed that the metabolic phenotype of the ketogenic diet (KD) is caused by the absence of carbohydrate and high fat content, however in rodents the protein content of KD affects weight gain and ketosis. In this study we examined the role of methionine and choline in mediating the metabolic effects of KD. We have found that choline was more effective than methionine in decreasing the liver steatosis of KD-fed mice. On the other hand, methionine supplementation was more effective than choline in restoring weight gain and normalizing the expression of several fatty acid and inflammatory genes in the liver of KD-fed mice. Our results indicate that choline and methionine restriction rather than carbohydrate restriction underlies many of the metabolic effects of KD.

  12. Epilepsy and the concentrations of plasma amino acids in humans.

    PubMed

    Huxtable, R J; Laird, H; Lippincott, S E; Walson, P

    1983-01-01

    We have examined the correlation between the presence of epilepsy in humans, and plasma amino acid levels. Subjects were divided into those having pure generalized tonic-clonic seizures (grand mal group), those having generalized tonic-clonic seizures plus other types of epilepsy (mixed group), and those suffering from epilepsies other than grand mal (no grand mal group). Compared to non-epileptic controls, the grand mal group had significantly higher fasting plasma levels of aspartate (100% increase) and glutamate (380% increase) but significant decreases were seen with phenylalanine (?23%), lysine (?27%), and tryptophan (?30%). The no grand mal group showed similar changes except for lysine. The mixed group showed elevations in glutamate, but decreases only in cysteine and methionine. In response to a high protein meal, plasma levels of alanine, cysteine and methionine rose significantly less for the no grand mal group compared to the control group. Increases in aspartate and glutamate concentrations strongly correlated with the prescription of phenytoin. However, the concentrations of these amino acids were not significantly correlated with the actual plasma levels of phenytoin.

  13. Molecular Basis for Lysine Specificity in the Yeast Ubiquitin-Conjugating Enzyme Cdc34 ▿

    PubMed Central

    Sadowski, Martin; Suryadinata, Randy; Lai, Xianning; Heierhorst, Jörg; Sarcevic, Boris

    2010-01-01

    Ubiquitin (Ub)-conjugating enzymes (E2s) and ubiquitin ligases (E3s) catalyze the attachment of Ub to lysine residues in substrates and Ub during monoubiquitination and polyubiquitination. Lysine selection is important for the generation of diverse substrate-Ub structures, which provides versatility to this pathway in the targeting of proteins to different fates. The mechanisms of lysine selection remain poorly understood, with previous studies suggesting that the ubiquitination site(s) is selected by the E2/E3-mediated positioning of a lysine(s) toward the E2/E3 active site. By studying the polyubiquitination of Sic1 by the E2 protein Cdc34 and the RING E3 Skp1/Cul1/F-box (SCF) protein, we now demonstrate that in addition to E2/E3-mediated positioning, proximal amino acids surrounding the lysine residues in Sic1 and Ub are critical for ubiquitination. This mechanism is linked to key residues composing the catalytic core of Cdc34 and independent of SCF. Changes to these core residues altered the lysine preference of Cdc34 and specified whether this enzyme monoubiquitinated or polyubiquitinated Sic1. These new findings indicate that compatibility between amino acids surrounding acceptor lysine residues and key amino acids in the catalytic core of ubiquitin-conjugating enzymes is an important mechanism for lysine selection during ubiquitination. PMID:20194622

  14. Structure-function validation of high lysine analogs of alpha-hordothionin designed by protein modeling.

    PubMed

    Rao, A G; Hassan, M; Hempel, J C

    1994-12-01

    Cereal grains and legume seeds, which are key protein sources for the vegetarian diet, are generally deficient in essential amino acids. Maize, in particular, is deficient in lysine. The inherent lack of lysine-rich proteins in maize has necessitated the search for heterologous proteins enriched in this amino acid, the isolation of the corresponding gene and its ultimate introduction into maize through plant transformation techniques. However, a rate-limiting step to this strategy has been the availability of plant-derived lysine-rich proteins. An appealing solution to the problem is to artificially increase the lysine content of a given protein by mutating appropriate residues to lysine. Here, we expound this strategy, starting with the protein alpha-hordothionin that is derived from barley seeds and consists of five lysine residues in a total of 45 amino acids (11% lysine). To facilitate rational substitutions, the 3-D structure of the protein has been determined by homology modeling with crambin. Based on this model, we have identified surface residues amenable to substitution with lysine. Furthermore, the acceptability of the mutations has been validated through the synthesis and characterization of the derivatives. To this end, our approach has permitted the creation of a modified alpha-hordothionin protein that has a lysine content of approximately 27% and retains the antifungal activity of the wild-type protein.

  15. Characterization of a second lysine decarboxylase isolated from Escherichia coli.

    PubMed Central

    Kikuchi, Y; Kojima, H; Tanaka, T; Takatsuka, Y; Kamio, Y

    1997-01-01

    We report here on the existence of a new gene for lysine decarboxylase in Escherichia coli K-12. The hybridization experiments with a cadA probe at low stringency showed that the homologous region of cadA was located in lambda Kohara phage clone 6F5 at 4.7 min on the E. coli chromosome. We cloned the 5.0-kb HindIII fragment of this phage clone and sequenced the homologous region of cadA. This region contained a 2,139-nucleotide open reading frame encoding a 713-amino-acid protein with a calculated molecular weight of 80,589. Overexpression of the protein and determination of its N-terminal amino acid sequence defined the translational start site of this gene. The deduced amino acid sequence showed 69.4% identity to that of lysine decarboxylase encoded by cadA at 93.7 min on the E. coli chromosome. In addition, the level of lysine decarboxylase activity increased in strains carrying multiple copies of the gene. Therefore, the gene encoding this lysine decarboxylase was designated Idc. Analysis of the lysine decarboxylase activity of strains containing cadA, ldc, or cadA ldc mutations indicated that ldc was weakly expressed under various conditions but is a functional gene in E. coli. PMID:9226257

  16. Purification and characterization of two human erythrocyte arylamidases preferentially hydrolysing N-terminal arginine or lysine residues.

    PubMed Central

    Mäkinen, K K; Mäkinen, P L

    1978-01-01

    Two arylamidases (I and II) were purified from human erythrocytes by a procedure that comprised removal of haemoglobin from disrupted cells with CM-Sephadex D-50, followed by treatment of the haemoglobin-free preparation subsequently with DEAE-cellulose, gel-permeation chromatography on Sephadex G-200, gradient solubilization on Celite, isoelectric focusing in a pH gradient from 4 to 6, gel-permeation chromatography on Sephadex G-100 (superfine), and finally affinity chromatography on Sepharose 4B covalently coupled to L-arginine. In preparative-scale purifications, enzymes I and II were separated at the second gel-permeation chromatography. Enzyme II was obtained as a homogeneous protein, as shown by several criteria. Enzyme I hydrolysed, with decreasing rates, the L-amino acid 2-naphtylamides of lysine, arginine, alanine, methionine, phenylalanine and leucine, and the reactions were slightly inhibited by 0.2 M-NaCl. Enzyme II hydrolysed most rapidly the corresponding derivatives of arginine, leucine, valine, methionine, proline and alanine, in that order, and the hydrolyses were strongly dependent on Cl-. The hydrolysis of these substrates proceeded rapidly at physiological Cl- concentration (0.15 M). The molecular weights (by gel filtration) of enzymes I and II were 85 000 and 52 500 respectively. The pH optimum was approx. 7.2 for both enzymes. The isoelectric point of enzyme II was approx. 4.8. Enzyme I was activated by Co2+, which did not affect enzyme II to any noticeable extent. The kinetics of reactions catalysed by enzyme I were characterized by strong substrate inhibition, but enzyme II was not inhibited by high substrate concentrations. The Cl- activated enzyme II also showed endopeptidase activity in hydrolysing bradykinin. PMID:743227

  17. Metabolic Fate of Cysteine and Methionine in Rumen Digesta

    PubMed Central

    Nader, C. J.; Walker, D. J.

    1970-01-01

    Estimates were obtained of the extent to which cysteine and methionine were incorporated into the protein of the microbes of rumen digesta without prior degradation and resynthesis. By using the amino acids labeled with both 35S and 14C, it was observed that a large proportion of the 35S appeared in the sulfide pool and of the 14C appeared in volatile fatty acids. By isolating the appropriate amino acid, obtaining the 14C to 35S ratio, and comparing this with the ratio in the added amino acid, the degree of direct incorporation was calculated. For cysteine it was estimated that at most 1% and for methionine, at most 11% of the amino acid in the free pool was incorporated unchanged into microbial protein. As a consequence of these findings, it is considered that the method for measuring microbial protein synthesis in rumen digesta based upon incorporation of 35S from the free sulfide pool is not seriously affected by direct utilization of sulfur amino acids arising from dietary sources. PMID:5485079

  18. Basis for the equilibrium constant in the interconversion of l-lysine and l-beta-lysine by lysine 2,3-aminomutase.

    PubMed

    Chen, Dawei; Tanem, Justinn; Frey, Perry A

    2007-02-01

    l-beta-lysine and beta-glutamate are produced by the actions of lysine 2,3-aminomutase and glutamate 2,3-aminomutase, respectively. The pK(a) values have been titrimetrically measured and are for l-beta-lysine: pK(1)=3.25 (carboxyl), pK(2)=9.30 (beta-aminium), and pK(3)=10.5 (epsilon-aminium). For beta-glutamate the values are pK(1)=3.13 (carboxyl), pK(2)=3.73 (carboxyl), and pK(3)=10.1 (beta-aminium). The equilibrium constants for reactions of 2,3-aminomutases favor the beta-isomers. The pH and temperature dependencies of K(eq) have been measured for the reaction of lysine 2,3-aminomutase to determine the basis for preferential formation of beta-lysine. The value of K(eq) (8.5 at 37 degrees C) is independent of pH between pH 6 and pH 11; ruling out differences in pK-values as the basis for the equilibrium constant. The K(eq)-value is temperature-dependent and ranges from 10.9 at 4 degrees C to 6.8 at 65 degrees C. The linear van't Hoff plot shows the reaction to be enthalpy-driven, with DeltaH degrees =-1.4 kcal mol(-1) and DeltaS degrees =-0.25 cal deg(-1) mol(-1). Exothermicity is attributed to the greater strength of the bond C(beta)-N(beta) in l-beta-lysine than C(alpha)-N(alpha) in l-lysine, and this should hold for other amino acids.

  19. Production of amino acids by free-living heterotrophic nitrogen-fixing bacteria.

    PubMed

    González-López, J; Martínez-Toledo, M V; Rodelas, B; Pozo, C; Salmerón, V

    1995-03-01

    Interest of microbial production of amino acids has been increased greatly since development of biotechnological methods. These methods represent a perspective way applied in a future large-scale manufacture of inexpensive amino acids. In this context, the isolation of producing organisms that may be exploited in the desing of alternative methods for the production of amino acids could be of primary importance.In this review we will describe the liberation of amino acids (methionine, lysine, arginine, tryptophane and glutamic acid) byAzotobacter andAzospirillum during growth in culture media with different carbon sources under diazotrophic and adiazotrophic conditions. These organisms may be useful in developing new methods for the industrial production of amino acids.

  20. l-lysine production by Bacillus methanolicus: Genome-based mutational analysis and l-lysine secretion engineering.

    PubMed

    Nærdal, Ingemar; Netzer, Roman; Irla, Marta; Krog, Anne; Heggeset, Tonje Marita Bjerkan; Wendisch, Volker F; Brautaset, Trygve

    2017-02-20

    Bacillus methanolicus is a methylotrophic bacterium with an increasing interest in academic research and for biotechnological applications. This bacterium was previously applied for methanol-based production of l-glutamate, l-lysine and the five-carbon diamine cadaverine by wild type, classical mutant and recombinant strains. The genomes of two different l-lysine secreting B. methanolicus classical mutant strains, NOA2#13A52-8A66 and M168-20, were sequenced. We focused on mutational mapping in genes present in l-lysine and other relevant amino acid biosynthetic pathways, as well as in the primary cell metabolism important for precursor supply. In addition to mutations in the aspartate pathway genes dapG, lysA and hom-1, new mutational target genes like alr, proA, proB1, leuC, odhA and pdhD were identified. Surprisingly, no mutations were found in the putative l-lysine transporter gene lysE(MGA3). Inspection of the wild type B. methanolicus strain PB1 genome sequence identified two homologous putative l-lysine transporter genes, lysE(PB1) and lysE2(PB1). The biological role of these putative l-lysine transporter genes, together with the heterologous l-lysine exporter gene lysE(Cg) from Corynebacterium glutamicum, were therefore investigated. Our results demonstrated that the titer of secreted l-lysine in B. methanolicus was significantly increased by overexpression of lysE(Cg) while overexpression of lysE(MGA3), lysE(PB1) and lysE2(PB1) had no measurable effect.

  1. Evaluation of apparent ileal digestibility of amino acids in Chinese corn by-products for growing-finishing pigs.

    PubMed

    Liang, Guo; Li, Defa; Wang, Fenglai; Dai, Jianguo; Yang, Wenjun

    2003-04-01

    Twelve crossbred barrows (BW 47 +/- 2.81 kg) were fitted with a T-cannula in the terminal ileum and fed one of ten diets containing corn gluten meal, dried distillers grain or dried distillers grain with solubles as the sole protein source, according to a 6 x 6 Latin Square design. Lysine, tryptophan and methionine were, the first, second and third limiting amino acids in these by-products. The ileal apparent digestibility of methionine, tyrosine, leucine and phenylalanine in corn gluten meal was high. The ileal apparent digestibility of tryptophan, tyrosine and leucine differed significantly in corn gluten meal. The ileal apparent digestibility of amino acids was higher in dried distillers grain with solubles than in dried distillers grain. To predict the ileal apparent digestibility of tryptophan, methionine and threonine linear regression equations were derived. The results suggested that processing technique can influence the ileal apparent digestibility ofamino acids in corn by-products. It was concluded that the direct technique cannot accurately determine the ileal apparent digestibility of tryptophan in dried distillers grain with solubles and dried distillers grain, due to its low content. The content of an amino acid in a feedstuff used as the sole source of protein must meet the minimum requirement of pigs when the ileal apparent digestibility of constituent amino acids is determined using the direct technique.

  2. Methionine oxidation by peroxymonocarbonate, a reactive oxygen species formed from CO2/bicarbonate and hydrogen peroxide.

    PubMed

    Richardson, David E; Regino, Celeste A S; Yao, Huirong; Johnson, Jodie V

    2003-12-15

    Kinetic and thermodynamic evidence is reported for the role of the peroxymonocarbonate ion, HCO4-, as a reactive oxygen species in biology. Peroxymonocarbonate results from the equilibrium reaction of hydrogen peroxide with bicarbonate via the perhydration of CO2. The kinetic parameters for HCO4- oxidation of free methionine have been obtained (k1 = 0.48 +/- 0.08 M(-1)s(-1) by a spectrophotometric initial rate method). At the physiological concentration of bicarbonate in blood ( approximately 25 mM), it is estimated that peroxymonocarbonate formed in equilibrium with hydrogen peroxide will oxidize methionine approximately 2-fold more rapidly than plasma H2O2 itself. As an example of methionine oxidation in proteins, the bicarbonate-catalyzed hydrogen peroxide oxidation of alpha1-proteinase inhibitor (alpha1-PI) has been investigated via its inhibitory effect on porcine pancreatic elastase activity. The second-order rate constant for HCO4- oxidation of alpha1-PI (0.36 +/- 0.06 M(-1)s(-1)) is comparable to that of free methionine, suggesting that methionine oxidation is occurring. Further evidence for methionine oxidation, specifically involving Met358 and Met351 of the alpha1-PI reactive center loop, has been obtained through amino acid analyses and mass spectroscopic analyses of proteolytic digests of the oxidized alpha1-PI. These results strongly suggest that HCO4- should be considered a reactive oxygen species in aerobic metabolism.

  3. Extended amino acid sequences around the active-site lysine residue of class-I fructose 1,6-bisphosphate aldolases from rabbit muscle, sturgeon muscle, trout muscle and ox liver.

    PubMed Central

    Benfield, P A; Forcina, B G; Gibbons, I; Perham, R N

    1979-01-01

    1. Amino acid sequences covering the region between residues 173 and 248 [adopting the numbering system proposed by Lai, Nakai & Chang (1974) Science 183, 1204-1206] were derived for trout (Salmo trutta) muscle aldolase and for ox liver aldolase. A comparable sequence was derived for residues 180-248 of sturgeon (Acipenser transmontanus) muscle aldolase. The close homology with the rabbit muscle enzyme was used to align the peptides of the other aldolases from which the sequences were derived. The results also allowed a partial sequence for the N-terminal 39 residues for the ox liver enzyme to be deduced. 2. In the light of the strong homology evinced for these enzymes, a re-investigation of the amino acid sequence of rabbit muscle aldolase between residues 181 and 185 was undertaken. This indicated the presence of a hitherto unsuspected -Ile-Val-sequence between residues 181 and 182 and the need to invert the sequence -Glu-Val- to -Val-Glx- at positions 184 and 185. 3. Comparison of the available amino acid sequences of these enzymes suggested an early evolutionary divergence of the genes for muscle and liver aldolases. It was also consistent with other evidence that the central region of the primary structure of these enzymes (which includes the active-site lysine-227) forms part of a conserved folding domain in the protein subunit. 4. Detailed evidence for the amino acid sequences proposed has been deposited as Suy Lending Division, Boston Spa, Wetherby, West Yorkshire LS23 7BQ, U.K., from whom copies can be obtained on the terms indicated in Biochem. J. (1978) 169, 5. PMID:534504

  4. Metabolic pathways and fermentative production of L-aspartate family amino acids.

    PubMed

    Park, Jin Hwan; Lee, Sang Yup

    2010-06-01

    The L-aspartate family amino acids (AFAAs), L-threonine, L-lysine, L-methionine and L-isoleucine have recently been of much interest due to their wide spectrum of applications including food additives, components of cosmetics and therapeutic agents, and animal feed additives. Among them, L-threonine, L-lysine and L-methionine are three major amino acids produced currently throughout the world. Recent advances in systems metabolic engineering, which combine various high-throughput omics technologies and computational analysis, are now facilitating development of microbial strains efficiently producing AFAAs. Thus, a thorough understanding of the metabolic and regulatory mechanisms of the biosynthesis of these amino acids is urgently needed for designing system-wide metabolic engineering strategies. Here we review the details of AFAA biosynthetic pathways, regulations involved, and export and transport systems, and provide general strategies for successful metabolic engineering along with relevant examples. Finally, perspectives of systems metabolic engineering for developing AFAA overproducers are suggested with selected exemplary studies.

  5. Influence of various nitrogen applications on protein and amino acid profiles of amaranth and quinoa.

    PubMed

    Thanapornpoonpong, Sa-nguansak; Vearasilp, Suchada; Pawelzik, Elke; Gorinstein, Shela

    2008-12-10

    The effect of nitrogen application levels (0.16 and 0.24 g N kg(-1) soil) on seed proteins and their amino acid compositions of amaranth (Amaranthus spp.) and quinoa (Chenopodium quinoa Willd) was studied. Total proteins of amaranth and quinoa had high contents of lysine (6.3-8.2 g 100 g(-1) protein) but low contents of methionine (1.2-1.8 g 100 g(-1) protein). Seed proteins were fractionated on the basis of different solubility in water, saline, and buffer as albumin-1 (Albu-1), albumin-2 (Albu-2), globulin (Glob), and glutelin (Glu) and were identified by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Albu-1 was high in lysine (5.4-8.6 g 100 g(-1) protein), while Albu-2, which is a part of storage proteins, had a high leucine content (7.2-8.9 g 100 g(-1) protein) as an effect of different nitrogen application levels. Glu fractions were well-balanced in their essential amino acids with the exception of methionine. In conclusion, nitrogen application can be used for the nutritional improvement in human diet by increasing and maintaining protein and essential amino acid contents.

  6. Crystallography captures catalytic steps in human methionine adenosyltransferase enzymes

    PubMed Central

    Murray, Ben; Antonyuk, Svetlana V.; Marina, Alberto; Lu, Shelly C.; Mato, Jose M.; Hasnain, S. Samar; Rojas, Adriana L.

    2016-01-01

    The principal methyl donor of the cell, S-adenosylmethionine (SAMe), is produced by the highly conserved family of methionine adenosyltranferases (MATs) via an ATP-driven process. These enzymes play an important role in the preservation of life, and their dysregulation has been tightly linked to liver and colon cancers. We present crystal structures of human MATα2 containing various bound ligands, providing a “structural movie” of the catalytic steps. High- to atomic-resolution structures reveal the structural elements of the enzyme involved in utilization of the substrates methionine and adenosine and in formation of the product SAMe. MAT enzymes are also able to produce S-adenosylethionine (SAE) from substrate ethionine. Ethionine, an S-ethyl analog of the amino acid methionine, is known to induce steatosis and pancreatitis. We show that SAE occupies the active site in a manner similar to SAMe, confirming that ethionine also uses the same catalytic site to form the product SAE. PMID:26858410

  7. Effects of intravenous infusion of amino acids and glucose on the yield and concentration of milk protein in dairy cows.

    PubMed

    Kim, C H; Kim, T G; Choung, J J; Chamberlain, D G

    2001-02-01

    To test the hypothesis that the availability of glucose or its precursors can influence the response of milk protein concentration to the intravenous infusion of amino acids, five cows were used in a 5 x 5 Latin square design with period lengths of 7 d. The five treatments were the basal diet of grass silage ad lib. plus 5 kg/d of a cereal-based supplement containing feather meal (Basal); Basal plus 4 g/d histidine, 8 g/d methionine and 26 g/d lysine (4H); Basal plus 8 g/d histidine, 8 g/d methionine and 26 g/d lysine (SH); and these two amino acid mixtures together with 600 g/d of gluctose (4HG and 8HG respectively). Earlier experiments with this basal diet had shown that histidine was first-limiting for secretion of milk protein, followed by methionine and lysine. The yield of milk protein was increased progressively with the amount of histidine infused. The efficiency of transfer of histidine into milk protein was 0.42 for the 4H and 4HG and 0.35 for the 8H and 8HG treatments, and the concentration of milk protein was increased over Basal by all infusion treatments. However, milk protein concentrations were higher, and lactose concentrations in the milk were lower, in the absence of added glucose. Concentrations of insulin in blood plasma were not affected by treatment. It is concluded that, with the treatments without added glucose, a shortage of glucose prevented an increase in lactose secretion, and hence limited the increase in milk yield, leading to an increased concentration of protein in the milk.

  8. Covalent modification of a highly reactive and essential lysine residue of ribulose-1,5-bisphosphate carboxylase/oxygenase activase.

    PubMed

    Salvucci, M E

    1993-10-01

    Chemical modification of ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco) activase with water-soluble N-hydroxysuccinimide esters was used to identify a reactive lysyl residue that is essential for activity. Incubation of Rubisco activase with sulfosuccinimidyl-7-amino-4-methylcoumarin-3-acetate (AMCA-sulfo-NHS) or sulfosuccinimidyl-acetate (sulfo-NHS-acetate) caused progressive inactivation of ATPase activity and concomitant loss of the ability to activate Rubisco. AMCA-sulfo-NHS was the more potent inactivator of Rubisco activase, exhibiting a second-order rate constant for inactivation of 239 M-1 s-1 compared to 21 M-1 s-1 for sulfo-NHS-acetate. Inactivation of enzyme activity by AMCA-sulfo-NHS correlated with the incorporation of 1.9 mol of AMCA per mol of 42-kD Rubisco activase monomer. ADP, a competitive inhibitor of Rubisco activase, afforded considerable protection against inactivation of Rubisco activase and decreased the amount of AMCA incorporated into the Rubisco activase monomer. Sequence analysis of the major labeled peptide from AMCA-sulfo-NHS-modified enzyme showed that the primary site of modification was lysine-247 (K247) in the tetrapeptide methionine-glutamic acid-lysine-phenylalanine. Upon complete inactivation of ATPase activity, modification of K247 accounted for 1 mol of AMCA incorporated per mol of Rubisco activase monomer. Photoaffinity labeling of AMCA-sulfo-NHS- and sulfo-NHS-acetate-modified Rubisco activase with ATP analogs derivatized on either the adenine base or on the gamma-phosphate showed that K247 is not essential for the binding of adenine nucleotides per se. Instead, the data indicated that the essentiality of K247 is probably due to an involvement of this highly reactive, species-invariant residue in an obligatory interaction that occurs between the protein and the nucleotide phosphate during catalysis.

  9. Changes in amino acids and lipids during embryogenesis of European lobster, Homarus gammarus (Crustacea: Decapoda).

    PubMed

    Rosa, R; Calado, R; Andrade, A M; Narciso, L; Nunes, M L

    2005-02-01

    We studied the amino acid and lipid dynamics during embryogenesis of Homarus gammarus. Major essential amino acids (EAA) in the last stage of embryonic development were arginine, lysine and leucine; major nonessential amino acids (NEAA) were glutamic acid, aspartic acid, valine and glycine. The highest percent of utilization occurred in respect to EAA (27.8%), mainly due to a significant decrease (p<0.05) of methionine (38.3%) and threonine (36.0%). NEAA also decreased significantly (p<0.05, 11.4%), namely serine (38.1%), tyrosine (26.4%) and glutamic acid (25.7%). In contrast, the free amino acid content increased significantly (p<0.05) during embryonic development, especially the free nonessential amino acids (FNEAA). In the last stage, the most abundant FNEAA were glycine, proline, alanine and taurine, and the major free essential amino acids (FEAA) were arginine, lysine and leucine. Lipid content decreased significantly (p<0.05) during embryonic development. A substantial decrease in all neutral lipid classes was observed (>80% of utilization). Major fatty acids were 16:0, 18:0, 18:1n-9, 18:2n-6, 18:3n-3, 20:5n-3 and 22:6n-3. Unsaturated (UFA) and saturated fatty acids (SFA) were used up at similar rates (76.5% and 76.3%, respectively). Within UFA, monounsaturates (MUFA) were consumed more than polyunsaturates (PUFA) (82.9% and 67.5%, respectively).

  10. Interaction between ambient temperature and supplementation of synthetic amino acids on performance and carcass parameters in commercial male turkeys.

    PubMed

    Veldkamp, T; Ferket, P R; Kwakkel, R P; Nixey, C; Noordhuizen, J P

    2000-10-01

    An experiment with male turkeys was conducted to test the hypothesis that turkey production performance responds positively to extra crystalline amino acid supplementation (lysine, methionine, and threonine) when subjected to a high ambient temperature regimen (HT) in the grower period. Two diets were formulated to provide lysine, methionine, and threonine concentrations that either 1) met the breeder recommendations or 2) contained 10% higher lysine and methionine concentrations from 22 to 134 d of age and 10% higher threonine concentration from 22 to 68 d of age. Both diets were fed at two temperatures (15 or 25 C) from 42 d of age onward. At 134 d of age, turkeys on the HT had generally lower BW than those on the low temperature regimen (LT). Up to 68 d of age and from 106 to 134 d of age, feed intake of turkeys on the HT was significantly lower than that of turkeys on the LT. Up to 42 d of age, feed conversion ratio (FCR) of turkeys on the HT were significantly lower than those of turkeys on the LT. Significant treatment interactions were observed from 22 to 41 d of age. Turkeys fed the amino acid-supplemented diets on the LT had significantly reduced FCR, whereas those on the HT did not respond. From 69 to 105 d of age, turkeys on the HT that were fed the supplemented diets had significantly increased FCR, but there were no dietary effects among turkeys on the LT. There were no consistent diet effects on growth performance or carcass yields. Breast meat yields of turkeys on the LT were higher (33.5 vs 32.1%), and drum yields were lower (12.7 vs 13.0%), than those of turkeys on the HT. There were no significant amino acid balance x ambient temperature effects on processing yields. The hypothesis of this experiment could be rejected as production performance did not respond positively to extra supplementation of lysine, methionine, and threonine when subjected to an HT.

  11. Molecular cloning and characterization of l-methionine γ-lyase from Streptomyces avermitilis.

    PubMed

    Kudou, Daizou; Yasuda, Eri; Hirai, Yoshiyuki; Tamura, Takashi; Inagaki, Kenji

    2015-10-01

    A pyridoxal 5'-phosphate-dependent methionine γ-lyase (MGL) was cloned from Streptomyces avermitilis catalyzed the degradation of methionine to α-ketobutyrate, methanethiol, and ammonia. The sav7062 gene (1,242 bp) was corresponded to 413 amino acid residues with a molecular mass of 42,994 Da. The deduced amino acid sequence showed a high degree of similarity to those of other MGL enzymes. The sav7062 gene was overexpressed in Escherichia coli. The enzyme was purified to homogeneity and exhibited the MGL catalytic activities. We cloned the enzyme that has the MGL activity in Streptomyces for the first time.

  12. Complexation des acides aminés basiques arginine, histidine et lysine avec l'ADN plasmidique en solution aqueuse : participation à la capture de radicaux sous irradiation X à 1,5 keV

    NASA Astrophysics Data System (ADS)

    Tariq Khalil, Talat; Taillefumier, Baptiste; Boulanouar, Omar; Mavon, Christophe; Fromm, Michel

    2016-09-01

    L'environnement chimique de l'ADN en situation biologique est complexe notam-ment en raison de la présence d'histones, protéines nucléaires, associées en quantité approximativement égales à l'ADN pour former la chromatine. Les histones possèdent de nombreux radicaux basiques arginine et lysine chargés positivement et dont la majorité se trouve sur les chaînes émergentes, l'ADN présente quant à lui des charges négatives sur ses groupements phosphates localisés tout au long de la double hélice. Dans cette étude, la complexité de la structure de la chromatine nucléaire est dans un premier temps mimée en solution aqueuse par la formation de complexes entre un ADN plasmidique sonde et les trois acides aminés basiques, Arg, His, Lys, qui, mis à part His, sont protonés au pH physiologique. Ces acides aminés libres en solution sont réputés être des capteurs efficaces de radicaux libres, notamment pour le radical hydroxyle, conférant ainsi un pouvoir protecteur vis-à-vis des effets indirects sur l'ADN en situation d'exposition aux rayonnements ionisants. A concentration fixée, les capacités de capture des acides aminés libres, σ, pour le radical hydroxyle sont typiquement les suivantes σHis ≈σArg > σLys (σLys ≈ 0,1 × σArg). Nous avons mesuré les taux de cassures simple brin par plasmide et par Gray (χ) lors d'expositions de solutions aqueuses de complexes [acide aminé - ADN plasmidique] aux rayons X ultra-mous (1,5 keV). A concentrations égales, les trois acides aminés complexés et présents en large excès ne manifestent pas une capacité de protection de l'ADN proportionnelle à leur capacité de capture libre et en solution ; on trouve en effet des taux de cassures dans l'ordre suivant χHis > χArg > χLys (χLys ≈ 0,01 χArg). Après avoir détaillé le mode opératoire de ces mesures, nous analyserons sur des bases bibliographiques, les modes spécifiques d'interaction des acides aminés basiques avec l'ADN. La sp

  13. Chemical composition and amino acid profiles of goose muscles from native Polish breeds.

    PubMed

    Okruszek, A; Woloszyn, J; Haraf, G; Orkusz, A; Werenska, M

    2013-04-01

    The aim of the study was to compare the chemical and amino acid composition of breast (pectoralis major) and thigh (biceps femoris) muscles in 17-wk-old geese from 2 Polish conservative flocks: Rypińska (Ry, n = 20) and Garbonosa (Ga, n = 20). The geese were fed ad libitum during the experimental period on the same complete feed. Genotypes affected the moisture and fat content of breast and thigh meat. The Ga geese were characterized by higher moisture as well as lower fat lipid content compared with the Ry breast and thigh muscles. The amino acid proportions of meat proteins depended on the goose flock and type of muscles, where significant differences were found. The proteins of Ga breast muscles contained more glutamic acid, glycine, lysine, tryptophan, histidine, and methionine, and less aspartic acid, proline, serine, leucine, valine, phenyloalanine, tyrosine, and threonine than the Ry geese (P ≤ 0.05). The proteins of Ry thigh muscles were characterized by higher content of proline, serine, and essential amino acids (without lysine and methionine) and lower glutamic and asparagine acid, alanine, and glycine compared with the Ga flock. According to the Food and Agriculture Organization of the United Nations/World Health Organization (1991) standard, tryptophan was the amino acid limiting the nutritional value of meat proteins of Ry breast muscles (amino acid score for tryptophan = 90%). Except for tryptophan, the meat proteins of the investigated raw materials contained more essential amino acids than the standard. The total content of essential amino acids for all investigated muscles was also higher (52.51 to 55.54%) than the standard (33.90%). It is evident that muscle protein from both flocks of geese have been characterized by high nutritional value. The values of the essential amino acid index of breast muscle proteins were similar in both flocks.

  14. Near-infrared reflectance spectroscopy (NIRS) enables the fast and accurate prediction of essential amino acid contents. 2. Results for wheat, barley, corn, triticale, wheat bran/middlings, rice bran, and sorghum.

    PubMed

    Fontaine, Johannes; Schirmer, Barbara; Hörr, Jutta

    2002-07-03

    Further NIRS calibrations were developed for the accurate and fast prediction of the total contents of methionine, cystine, lysine, threonine, tryptophan, and other essential amino acids, protein, and moisture in the most important cereals and brans or middlings for animal feed production. More than 1100 samples of global origin collected over five years were analyzed for amino acids following the Official Methods of the United States and European Union. Detailed data and graphics are given to characterize the obtained calibration equations. NIRS was validated with 98 independent samples for wheat and 78 samples for corn and compared to amino acid predictions using linear crude protein regression equations. With a few exceptions, validation showed that 70-98% of the amino acid variance in the samples could be explained using NIRS. Especially for lysine and methionine, the most limiting amino acids for farm animals, NIRS can predict contents in cereals much better than crude protein regressions. Through low cost and high speed of analysis NIRS enables the amino acid analysis of many samples in order to improve the accuracy of feed formulation and obtain better quality and lower production costs.

  15. Exploration of the binding modes of buffalo PGRP1 receptor complexed with meso-diaminopimelic acid and lysine-type peptidoglycans by molecular dynamics simulation and free energy calculation.

    PubMed

    Sahoo, Bikash Ranjan; Dubey, Praveen Kumar; Goyal, Shubham; Bhoi, Gopal Krushna; Lenka, Santosh Kumar; Maharana, Jitendra; Pradhan, Sukanta Kumar; Kataria, Ranjit Singh

    2014-09-05

    The peptidoglycan recognition proteins (PGRPs) are the key components of innate-immunity, and are highly specific for the recognition of bacterial peptidoglycans (PGN). Among different mammalian PGRPs, the PGRP1 binds to murein PGN of Gram-positive bacteria (lysine-type) and also have bactericidal activity towards Gram-negative bacteria (diaminopimelic acid or Dap-type). Buffaloes are the major sources of milk and meat in Asian sub-continents and are highly exposed to bacterial infections. The PGRP activates the innate-immune signaling, but their studies has been confined to limited species due to lack of structural and functional information. So, to understand the structural constituents, 3D model of buffalo PGRP1 (bfPGRP1) was constructed and conformational and dynamics properties of bfPGRP1 was studied. The bfPGRP1 model highly resembled human and camel PGRP structure, and shared a highly flexible N-terminus and centrally placed L-shaped cleft. Docking simulation of muramyl-tripeptide, tetrapeptide, pentapeptide-Dap-(MTP-Dap, MTrP-Dap and MPP-Dap) and lysine-type (MTP-Lys, MTrP-Lys and MPP-Lys) in AutoDock 4.2 and ArgusLab 4.0.1 anticipated β1, α2, α4, β4, and loops connecting β1-α2, α2-β2, β3-β4 and α4-α5 as the key interacting domains. The bfPGRP1-ligand complex molecular dynamics simulation followed by free binding energy (BE) computation conceded BE values of -18.30, -35.53, -41.80, -25.03, -24.62 and -22.30 kJ mol(-1) for MTP-Dap, MTrP-Dap, MPP-Dap, MTP-Lys, MTrP-Lys and MPP-Lys, respectively. The groove-surface and key binding residues involved in PGN-Dap and Lys-type interaction intended by the molecular docking, and were also accompanied by significant BE values directed their importance in pharmacogenomics, and warrants further in vivo studies for drug targeting and immune signaling pathways exploration.

  16. Conversion of Methionine to Thiols by Lactococci, Lactobacilli, and Brevibacteria†

    PubMed Central

    Dias, Benjamin; Weimer, Bart

    1998-01-01

    Methanethiol has been strongly associated with desirable Cheddar cheese flavor and can be formed from the degradation of methionine (Met) via a number of microbial enzymes. Methionine γ-lyase is thought to play a major role in the catabolism of Met and generation of methanethiol in several species of bacteria. Other enzymes that have been reported to be capable of producing methanethiol from Met in lactic acid bacteria include cystathionine β-lyase and cystathionine γ-lyase. The objective of this study was to determine the production, stability, and activities of the enzymes involved in methanethiol generation in bacteria associated with cheese making. Lactococci and lactobacilli were observed to contain high levels of enzymes that acted primarily on cystathionine. Enzyme activity was dependent on the concentration of sulfur amino acids in the growth medium. Met aminotransferase activity was detected in all of the lactic acid bacteria tested and α-ketoglutarate was used as the amino group acceptor. In Lactococcus lactis subsp. cremoris S2, Met aminotransferase was repressed with increasing concentrations of Met in the growth medium. While no Met aminotransferase activity was detected in Brevibacterium linens BL2, it possessed high levels of l-methionine γ-lyase that was induced by addition of Met to the growth medium. Met demethiolation activity at pH 5.2 with 4% NaCl was not detected in cell extracts but was detected in whole cells. These data suggest that Met degradation in Cheddar cheese will depend on the organism used in production, the amount of enzyme released during aging, and the amount of Met in the matrix. PMID:9726877

  17. Sugar Substrates for l-Lysine Fermentation by Ustilago maydis

    PubMed Central

    Sánchez-Marroquín, A.; Ledezma, M.; Carreño, R.

    1970-01-01

    The extracellular production of l-lysine in media with cane sugar, blackstrap molasses, or clarified sugar-cane juice by a previously obtained mutant of Ustilago maydis was studied. Enzymatically inverted clarified juice (medium J-3) gave 2.9 g of lysine per liter under the following conditions: inoculum, 5%; pH 5.8; temperature, 30 C; KLa in the fermentors, 0.41 mmoles of O2 per liter per min; fermentation time, 72 hr. The concentrate, obtained by direct evaporation and drying of the fermentation broth, could be used as a possible feed supplement because of its amino-acid and vitamin content. PMID:5485081

  18. Conserved methionine dictates substrate preference in Nramp-family divalent metal transporters.

    PubMed

    Bozzi, Aaron T; Bane, Lukas B; Weihofen, Wilhelm A; McCabe, Anne L; Singharoy, Abhishek; Chipot, Christophe J; Schulten, Klaus; Gaudet, Rachelle

    2016-09-13

    Natural resistance-associated macrophage protein (Nramp) family transporters catalyze uptake of essential divalent transition metals like iron and manganese. To discriminate against abundant competitors, the Nramp metal-binding site should favor softer transition metals, which interact either covalently or ionically with coordinating molecules, over hard calcium and magnesium, which interact mainly ionically. The metal-binding site contains an unusual, but conserved, methionine, and its sulfur coordinates transition metal substrates, suggesting a vital role in their transport. Using a bacterial Nramp model system, we show that, surprisingly, this conserved methionine is dispensable for transport of the physiological manganese substrate and similar divalents iron and cobalt, with several small amino acid replacements still enabling robust uptake. Moreover, the methionine sulfur's presence makes the toxic metal cadmium a preferred substrate. However, a methionine-to-alanine substitution enables transport of calcium and magnesium. Thus, the putative evolutionary pressure to maintain the Nramp metal-binding methionine likely exists because it-more effectively than any other amino acid-increases selectivity for low-abundance transition metal transport in the presence of high-abundance divalents like calcium and magnesium.

  19. Methionine deprivation suppresses triple-negative breast cancer metastasis in vitro and in vivo

    PubMed Central

    Jang, Young Jin; Son, Joe Eun; Kwon, Jung Yeon; Lim, Tae-gyu; Kim, Sunghoon; Park, Jung Han Yoon; Kim, Jong-Eun; Lee, Ki Won

    2016-01-01

    Nutrient deprivation strategies have been proposed as an adjuvant therapy for cancer cells due to their increased metabolic demand. We examined the specific inhibitory effects of amino acid deprivation on the metastatic phenotypes of the human triple-negative breast cancer (TNBC) cell lines MDA-MB-231 and Hs 578T, as well as the orthotopic 4T1 mouse TNBC tumor model. Among the 10 essential amino acids tested, methionine deprivation elicited the strongest inhibitory effects on the migration and invasion of these cancer cells. Methionine deprivation reduced the phosphorylation of focal adhesion kinase, as well as the activity and mRNA expression of matrix metalloproteinases MMP-2 and MMP-9, two major markers of metastasis, while increasing the mRNA expression of tissue inhibitor of metalloproteinase 1 in MDA-MB-231 cells. Furthermore, methionine restriction downregulated the metastasis-related factor urokinase plasminogen activatior and upregulated plasminogen activator inhibitor 1 mRNA expression. Animals on the methionine-deprived diet showed lower lung metastasis rates compared to mice on the control diet. Taken together, these results suggest that methionine restriction could provide a potential nutritional strategy for more effective cancer therapy. PMID:27579534

  20. Formation of S-(carboxymethyl)-cysteine in rat liver mitochondrial proteins: effects of caloric and methionine restriction.

    PubMed

    Naudí, Alba; Jové, Mariona; Cacabelos, Daniel; Ayala, Victoria; Cabre, Rosanna; Caro, Pilar; Gomez, José; Portero-Otín, Manuel; Barja, Gustavo; Pamplona, Reinald

    2013-02-01

    Maillard reaction contributes to the chemical modification and cross-linking of proteins. This process plays a significant role in the aging process and determination of animal longevity. Oxidative conditions promote the Maillard reaction. Mitochondria are the primary site of oxidants due to the reactive molecular species production. Mitochondrial proteome cysteine residues are targets of oxidative attack due to their specific chemistry and localization. Their chemical, non-enzymatic modification leads to dysfunctional proteins, which entail cellular senescence and organismal aging. Previous studies have consistently shown that caloric and methionine restrictions, nutritional interventions that increase longevity, decrease the rate of mitochondrial oxidant production and the physiological steady-state levels of markers of oxidative damage to macromolecules. In this scenario, we have detected S-(carboxymethyl)-cysteine (CMC) as a new irreversible chemical modification in mitochondrial proteins. CMC content in mitochondrial proteins significantly correlated with that of the lysine-derived analog N (ε)-(carboxymethyl)-lysine. The concentration of CMC is, however, one order of magnitude lower compared with CML likely due in part to the lower content of cysteine with respect to lysine of the mitochondrial proteome. CMC concentrations decreases in liver mitochondrial proteins of rats subjected to 8.5 and 25 % caloric restriction, as well as in 40 and 80 % methionine restriction. This is associated with a concomitant and significant increase in the protein content of sulfhydryl groups. Data presented here evidence that CMC, a marker of Cys-AGE formation, could be candidate as a biomarker of mitochondrial damage during aging.

  1. Amino acid composition of some Amaranthus sp. grain proteins and of its fractions.

    PubMed

    Correa, A D; Jokl, L; Carlsson, R

    1986-09-01

    This study was carried out to determine the protein content of several Amaranthus sp. grains. Findings revealed this has a high lysine (5.3 to 6.3 of the protein) and sulphur amino acids content (3.4-4.0%), while leucine could well be limiting when those seeds are used as a sole protein source in food. Using the correction for in vitro protein digestibility, the chemical score varied from 50 to 67. The calculated protein efficiency ratios and biological values ranged from 1.39 to 1.80 and 53 to 68, respectively. Considering that amaranth grain is a good supplement to cereal grain, the protein of A. hypochondriacus HH5 (yellow seeds) and A. anclancalius (black seeds) was fractionated into albumin, globulin, prolamin and glutelin. The average proportions between those soluble proteins were 65:17:11:7, respectively. Albumin had the highest lysine content (7.3-8.2%), and globulin the highest methionine (4.1-5.3%) and phenylalanine (6.0-6.1%) content. Prolamin had the highest threonine (4.6-5.4%) and leucine (6.8-6.9%) content, while glutelin had a very low methionine content (0.6-1.0%). Based on the above-mentioned findings, the authors conclude the variation in the amino acid composition of the protein fractions can be used for genetic protein improvement.

  2. Effects of lysine-induced acute renal failure in dogs.

    PubMed

    Asanuma, Kentaro; Adachi, Kenji; Sugimoto, Tetsuro; Chiba, Shuichi

    2006-05-01

    This study investigates the effects of lysine-induced acute renal failure. Female dogs received a lysine hydrochloride (lysine) of 4500 mg/kg/day (3.75 ml/kg/hr) for 3 consecutive days. The dogs were observed for clinical signs. Body weights were recorded, food consumption and water consumption calculated, and urinalysis and blood biochemistry were performed daily. Plasma samples for amino acid determinations were obtained from all dogs, which were necropsied on Day 3. Histopathological examinations were done on all test animals. Compound-related findings include the following. Blood biochemistry results showed increases in ammonia, blood urea nitrogen, blood urea nitrogen/creatinine ratio, and creatinine. Urinary changes consisted of increases in urine volume, total protein, albumin, gamma-glutamyl transpeptidase, and N-acetyl-beta-D-glucosaminidase. In addition, macroscopic findings consisted of pale, congested capsule; microscopic findings consisted of hypertrophy of proximal convoluted tubule (mainly S1 segment), and degeneration/desquamation of urinary tubule (mainly S3 segment with hyaline casts) in the kidney. From these findings, it can be concluded that lysine is nephrotoxic in dogs. Nephrotoxicity of lysine may relate to direct tubular toxicity and to tubular obstruction.

  3. Site-saturation engineering of lysine 47 in cyclodextrin glycosyltransferase from Paenibacillus macerans to enhance substrate specificity towards maltodextrin for enzymatic synthesis of 2-O-D-glucopyranosyl-L-ascorbic acid (AA-2G).

    PubMed

    Han, Ruizhi; Liu, Long; Shin, Hyun-dong; Chen, Rachel R; Du, Guocheng; Chen, Jian

    2013-07-01

    In this work, the site-saturation engineering of lysine 47 in cyclodextrin glycosyltransferase (CGTase) from Paenibacillus macerans was conducted to improve the specificity of CGTase towards maltodextrin, which can be used as a cheap and easily soluble glycosyl donor for the enzymatic synthesis of 2-O-D-glucopyranosyl-L-ascorbic acid (AA-2G) by CGTase. When using maltodextrin as glycosyl donor, four mutants K47F (lysine→ phenylalanine), K47L (lysine→ leucine), K47V (lysine→ valine) and K47W (lysine→ tryptophan) showed higher AA-2G yield as compared with that produced by the wild-type CGTase. The transformation conditions (temperature, pH and the mass ratio of L-ascorbic acid to maltodextrin) were optimized and the highest titer of AA-2G produced by the mutant K47L could reach 1.97 g/l, which was 64.2% higher than that (1.20 g/l) produced by the wild-type CGTase. The reaction kinetics analysis confirmed the enhanced maltodextrin specificity, and it was also found that compared with the wild-type CGTase, the four mutants had relatively lower cyclization activities and higher disproportionation activities, which was favorable for AA-2G synthesis. The mechanism responsible for the enhanced substrate specificity was further explored by structure modeling and it was indicated that the enhancement of maltodextrin specificity may be due to the short residue chain and the removal of hydrogen bonding interactions between the side chain of residue 47 and the sugar at -3 subsite. Here the obtained mutant CGTases, especially the K47L, has a great potential in the production of AA-2G with maltodextrin as a cheap and easily soluble substrate.

  4. Plasma amino acid response to graded levels of escape protein.

    PubMed

    Gibb, D J; Klopfenstein, T J; Britton, R A; Lewis, A J

    1992-09-01

    A trial was conducted to examine the potential of using plasma amino acid responses to graded levels of escape protein to determine limiting amino acids in cattle. Growing calves (n = 120; mean BW = 220 +/- 21 kg) were fed a basal diet of corncob:sorghum silage (61:39) and were individually supplemented with distillers' dried grains (DDG), heat-damaged DDG (H-DDG), feather meal (FTH), or urea. The urea supplement was mixed with DDG and H-DDG to allow 0, 20, 35, 50, 65, or 80% of the supplemental CP to come from distillers' protein and maintain an 11.5% CP diet. Urea supplement was mixed with FTH to allow 0, 22, 39, 56, 73, or 90% of the supplemental CP to come from FTH. Dietary CP ranged from 11.5% at the 0% level to 17.3% at the 90% level. Plasma concentration of most essential plasma amino acids responded (P less than .10) linearly and(or) quadratically to increased escape protein. The broken-line response of plasma methionine at low DDG intake suggested that methionine was limiting at low levels of escape protein. An initial decrease followed by a plateau fit by a broken line indicated that histidine became limiting in FTH diets, and lysine eventually became limiting for DDG, H-DDG, and FTH diets before maximum BW gain was reached. Results indicate that plasma amino acid responses may identify amino acids that become limiting with increasing escape protein.

  5. Evaluation of Methionine Content in a High-Fat and Choline-Deficient Diet on Body Weight Gain and the Development of Non-Alcoholic Steatohepatitis in Mice

    PubMed Central

    Chiba, Tsuyoshi; Suzuki, Sachina; Sato, Yoko; Itoh, Tatsuki; Umegaki, Keizo

    2016-01-01

    Aim Non-alcoholic steatohepatitis (NASH) is a globally recognized liver disease. A methionine- and choline-deficient diet is used to induce NASH in mice; however, this diet also causes severe body weight loss. To resolve this issue, we examined the effects of methionine content in a high-fat and choline-deficient (HFCD) diet on body weight and the development of NASH in mice. Methods C57BL/6J mice (male, 10 weeks of age) were fed an L-amino acid rodent (control) diet, high-fat (HF) diet, or HFCD diet containing various amounts of methionine (0.1–0.6% (w/w)) for 12 weeks. Plasma lipid levels, hepatic lipid content and inflammatory marker gene expression were measured, and a pathological analysis was conducted to evaluate NASH. Results The 0.1% methionine in HFCD diet suppressed body weight gain, which was lower than that with control diet. On the other hand, the 0.2% methionine in HFCD diet yielded similar body weight gains as the control diet, while more than 0.4% methionine showed the same body weight gains as the HF diet. Liver weights and hepatic lipid contents were the greatest with 0.1% methionine and decreased in a methionine dose-dependent manner. Pathological analysis, NAFLD activity scores and gene expression levels in the liver revealed that 0.1% and 0.2% methionine for 12 weeks induced NASH, whereas 0.4% and 0.6% methionine attenuated the induction of NASH by HFCD diet. However, the 0.2% methionine in HFCD diet did not induce insulin resistance, despite the body weight gain. Conclusions The 0.2% methionine in HFCD diet for 12 weeks was able to induce NASH without weight loss. PMID:27723801

  6. Formation and inhibition of Nε-(carboxymethyl)lysine in saccharide-lysine model systems during microwave heating.

    PubMed

    Li, Lin; Han, Lipeng; Fu, Quanyi; Li, Yuting; Liang, Zhili; Su, Jianyu; Li, Bing

    2012-10-31

    N(ε)-(carboxymethyl) lysine (CML) is the most abundant advanced glycation end product (AGE), and frequently selected as an AGEs marker in laboratory studies. In this paper, the formation and inhibition of N(ε)-(carboxymethyl)lysine in saccharide-lysine model systems during microwave heating have been studied. The microwave heating treatment significantly promoted the formation of CML during Maillard reactions, which was related to the reaction temperature, time and type of saccharide. The order of CML formation for different saccharides was lactose > glucose > sucrose. Then, the inhibition effect on CML by five inhibitors was further examined. According to the results, ascorbic acid and tocopherol did not affect inhibition of CML, in contrast, thiamin, rutin and quercetin inhibited CML formation, and the inhibitory effects were concentration dependent.

  7. Functional Consequences of Methionine Oxidation of hERG Potassium Channels

    PubMed Central

    Su, Zhi; Limberis, James; Martin, Ruth L.; Xu, Rong; Kolbe, Katrin; Heinemann, Stefan H.; Hoshi, Toshinori; Cox, Bryan F.; Gintant, Gary A.

    2010-01-01

    Reactive species oxidatively modify numerous proteins including ion channels. Oxidative sensitivity of ion channels is often conferred by amino acids containing sulfur atoms, such as cysteine and methionine. Functional consequences of oxidative modification of methionine in hERG1 (human ether à go-go related gene 1), which encodes cardiac IKr channels, are unknown. Here we used chloramine-T (ChT), which preferentially oxidizes methionine, to examine the functional consequences of methionine oxidation of hERG channels stably expressed in a human embryonic kidney cell line (HEK 293) and native hERG channels in a human neuroblastoma cell line (SH-SY5Y). ChT (300 µM) significantly decreased whole-cell hERG current in both HEK 293 and SH-SY5Y cells. In HEK 293 cells, the effects of ChT on hERG current were time- and concentration-dependent, and were markedly attenuated in the presence of enzyme methionine sulfoxide reductase A that specifically repairs oxidized methionine. After treatment with ChT, the channel deactivation upon repolarization to −60 or −100 mV was significantly accelerated. The effect of ChT on channel activation kinetics was voltage-dependent; activation slowed during depolarization to +30 mV but accelerated during depolarization to 0 or −10 mV. In contrast, the reversal potential, inactivation kinetics, and voltage-dependence of steady-state inactivation remained unaltered. Our results demonstrate that the redox status of methionine is an important modulator of hERG channel. PMID:17624316

  8. Evidence for distinct L-methionine catabolic pathways in the yeast Geotrichum candidum and the bacterium Brevibacterium linens.

    PubMed

    Arfi, Kenza; Landaud, Sophie; Bonnarme, Pascal

    2006-03-01

    Tracing experiments were carried out to identify volatile and nonvolatile L-methionine degradation intermediates and end products in the yeast Geotrichum candidum and in the bacterium Brevibacterium linens, both of which are present in the surface flora of certain soft cheeses and contribute to the ripening reactions. Since the acid-sensitive bacterium B. linens is known to produce larger amounts and a greater variety of volatile sulfur compounds (VSCs) than the yeast G. candidum produces, we examined whether the L-methionine degradation routes of these microorganisms differ. In both microorganisms, methanethiol and alpha-ketobutyrate are generated; the former compound is the precursor of other VSCs, and the latter is subsequently degraded to 2,3-pentanedione, which has not been described previously as an end product of L-methionine catabolism. However, the L-methionine degradation pathways differ in the first steps of L-methionine degradation. L-Methionine degradation is initiated by a one-step degradation process in the bacterium B. linens, whereas a two-step degradation pathway with 4-methylthio-2-oxobutyric acid (MOBA) and 4-methylthio-2-hydroxybutyric acid (MHBA) as intermediates is used in the yeast G. candidum. Since G. candidum develops earlier than B. linens during the ripening process, MOBA and MHBA generated by G.candidum could also be used as precursors for VSC production by B. linens.

  9. Light-activated amino acid transport in Halobacterium halobium envelope vesicles

    NASA Technical Reports Server (NTRS)

    Macdonald, R. E.; Lanyi, J. K.

    1977-01-01

    Vesicles prepared from Halobacterium halobium cell envelopes accumulate amino acids in response to light-induced electrical and chemical gradients. Nineteen of 20 commonly occurring amino acids have been shown to be actively accumulated by these vesicles in response to illumination or in response to an artificially created Na+ gradient. On the basis of shared common carriers the transport systems can be divided into eight classes, each responsible for the transport of one or several amino acids: arginine, lysine, histidine; asparagine, glutamine; alanine, glycine, threonine, serine; leucine, valine, isoleucine, methionine; phenylalanine, tyrosine, tryptophan; aspartate; glutamate; proline. Available evidence suggests that these carriers are symmetrical in that amino acids can be transported equally well in both directions across the vesicle membranes. A tentative working model to account for these observations is presented.

  10. Evaluation of microwave-accelerated residue-specific acid cleavage for proteomic applications.

    PubMed

    Swatkoski, Stephen; Gutierrez, Peter; Wynne, Colin; Petrov, Alexey; Dinman, Jonathan D; Edwards, Nathan; Fenselau, Catherine

    2008-02-01

    Microwave-accelerated proteolysis using acetic acid has been shown to occur specifically on either or both sides of aspartic acid residues. This chemical cleavage has been applied to ovalbumin and several model peptides to test the effect on some of the more common post-translational modifications. No oxidation of methionine or cysteine was observed; however, hydrolysis of phosphate groups proceeds at a detectable rate. Acid cleavage was also extended to the yeast ribosome model proteome, where it provided information on 74% of that proteome. Aspartic acid occurs across the proteome with approximately half the frequency of the combined occurrence of the trypsin residues lysine and arginine, and implications of this are considered.

  11. Role of arginine 180 and glutamic acid 177 of ricin toxin A chain in enzymatic inactivation of ribosomes.

    PubMed Central

    Frankel, A; Welsh, P; Richardson, J; Robertus, J D

    1990-01-01

    The gene for ricin toxin A chain was modified by site-specific mutagenesis to change arginine 180 to alanine, glutamine, methionine, lysine, or histidine. Separately, glutamic acid 177 was changed to alanine and glutamic acid 208 was changed to aspartic acid. Both the wild-type and mutant proteins were expressed in Escherichia coli and, when soluble, purified and tested quantitatively for enzyme activity. A positive charge at position 180 was found necessary for solubility of the protein and for enzyme activity. Similarly, a negative charge with a proper geometry in the vicinity of position 177 was critical for ricin toxin A chain catalysis. When glutamic acid 177 was converted to alanine, nearby glutamic acid 208 could largely substitute for it. This observation provided valuable structural information concerning the nature of second-site mutations. Images PMID:1978925

  12. A GC-ECD method for estimation of free and bound amino acids, gamma-aminobutyric acid, salicylic acid, and acetyl salicylic acid from Solanum lycopersicum (L.).

    PubMed

    Meher, Hari Charan; Gajbhiye, Vijay T; Singh, Ghanendra

    2011-01-01

    A gas chromatograph with electron capture detection method for estimation of selected metabolites--amino acids (free and bound), gamma-aminobutyric acid (GABA), salicylic acid (SA), and acetyl salicylic acid (ASA) from tomato--is reported. The method is based on nitrophenylation of the metabolites by 1-fluoro-2, 4-dinitrobenzene under aqueous alkaline conditions to form dinitophenyl derivatives. The derivatives were stable under the operating conditions of GC. Analysis of bound amino acids comprised perchloric acid precipitation of protein, alkylation (carboxymethylation) with iodoacetic acid, vapor-phase hydrolysis, and derivatization with 1-fluoro-2,4-dinitrobenzene in that order. The metabolites were resolved in 35 min, using a temperature-programmed run. The method is rapid, sensitive, and precise. It easily measured the typical amino acids (aspartate, asparagine, glutamate, glutamine, alanine, leucine, lysine, and phenylalanine) used for identification and quantification of a protein, resolved amino acids of the same mass (leucine and isoleucine), satisfactorily measured sulfur amino acid (methionine, cystine, and cysteine), and quantified GABA, SA, and ASA, as well. The developed method was validated for specificity, linearity, and precision. It has been applied and recommended for estimation of 25 metabolites from Solanum lycopersicum (L.).

  13. Characterization of methionine oxidation and methionine sulfoxide reduction using methionine-rich cysteine-free proteins

    PubMed Central

    2012-01-01

    Background Methionine (Met) residues in proteins can be readily oxidized by reactive oxygen species to Met sulfoxide (MetO). MetO is a promising physiological marker of oxidative stress and its inefficient repair by MetO reductases (Msrs) has been linked to neurodegeneration and aging. Conventional methods of assaying MetO formation and reduction rely on chromatographic or mass spectrometry procedures, but the use of Met-rich proteins (MRPs) may offer a more streamlined alternative. Results We carried out a computational search of completely sequenced genomes for MRPs deficient in cysteine (Cys) residues and identified several proteins containing 20% or more Met residues. We used these MRPs to examine Met oxidation and MetO reduction by in-gel shift assays and immunoblot assays with antibodies generated against various oxidized MRPs. The oxidation of Cys-free MRPs by hydrogen peroxide could be conveniently monitored by SDS-PAGE and was specific for Met, as evidenced by quantitative reduction of these proteins with Msrs in DTT- and thioredoxin-dependent assays. We found that hypochlorite was especially efficient in oxidizing MRPs. Finally, we further developed a procedure wherein antibodies made against oxidized MRPs were isolated on affinity resins containing same or other oxidized or reduced MRPs. This procedure yielded reagents specific for MetO in these proteins, but proved to be ineffective in developing antibodies with broad MetO specificity. Conclusion Our data show that MRPs provide a convenient tool for characterization of Met oxidation, MetO reduction and Msr activities, and could be used for various aspects of redox biology involving reversible Met oxidation. PMID:23088625

  14. The importance of transmethylation reactions to methionine metabolism in sheep: effects of supplementation with creatine and choline.

    PubMed

    Lobley, G E; Connell, A; Revell, D

    1996-01-01

    The influence of administering the methylated products choline and creatine on methionine irreversible-loss rate (ILR) and recycling from homocysteine has been investigated in sheep fed close to energy and N equilibrium. Two methods to estimate methionine recycling were compared. The first involved [U-13C]methionine infused as part of a labelled amino acid mixture obtained from hydrolysed algal protein. In this approach the isotope dilution of methionine with all five C atoms labelled (m + 5) will represent the ILR which does not recycle through homocysteine, while that which includes molecules with C-1-C-4 labelled will allow for loss of the labelled methyl (5)-C atom and replacement by an unlabelled moiety in the remethylation of homocysteine. The second method involved a combined infusion of [1-13C]- and [S-methyl-2H3]methionine. These two approaches gave similar data for methionine ILR which does not include label recycled to the amino acid from homocysteine but differed for recycled methionine fluxes. Consequently the two procedures differed in the calculated extent of homocysteine methylation under control conditions (6 v. 28%). These extents of remethylation are within the range observed for the fed human subject, despite the fact that fewer dietary methyl groups are available for the ruminant. Using combined data from the infusions, significant depression of methionine recycling occurred in blood (P < 0.05), with a similar trend for plasma (P = 0.077), when choline plus creatine were infused. Wool growth, assessed by intradermal injection of [35S]cysteine, was not altered by supplementation with the methylated products. From changes in the label pattern of free methionine in aortal, hepatic portal and hepatic venous blood during U-13C-labelled algal hydrolysate infusion, the major sites of homocysteine remethylation appear to be the portal-drained viscera and the liver. This was confirmed by analysis of free methionine enrichments in various tissues

  15. Evaluating a quantitative methionine requirement for juvenile Pacific white shrimp Litopenaeus vannamei

    Technology Transfer Automated Retrieval System (TEKTRAN)

    A 10-wk feeding trial was conducted as a third study (all conducted in our laboratory) to determine a quantitative requirement of juvenile Litopenaeus vannamei for sulfur amino acid methionine. Juvenile shrimp (mean weight 0.61 +/- 0.13 g) were reared in 110-L aquaria in a seawater recirculating sy...

  16. Isotopic Dilution GC/MS Method for Methionine Determination in Biological Media

    NASA Astrophysics Data System (ADS)

    Horj, Elena; Iordache, Andreea; Culea, Monica

    2011-10-01

    The isotopic dilution mass spectrometry technique is the method of choice for sensitive and accurate determination of analytes in biological samples. The aim of this work was to establish a sensitive analytical method for the determination of methionine in different biological media. Quantitation of methionine from the resultant tracer spectrum requires deconvolution of the enrichment of the isotopomers. Deconvolution of the ion abundance ratios to yield tracer-to-tracee ratio for the isotopomer was done using Brauman's least squares approach. Comparison with regression curve calculation method is presented. The method was applied for amino-acids determination in beef, pork and fish meat.

  17. Effects of rumen-undegradable protein sources and supplemental 2-hydroxy-4-(methylthio)-butanoic acid and lysine-HCl on lactation performance in dairy cows.

    PubMed

    Johnson-VanWieringen, L M; Harrison, J H; Davidson, D; Swift, M L; von Keyserlingk, M A G; Vazquez-Anon, M; Wright, D; Chalupa, W

    2007-11-01

    One hundred primiparous and multiparous Holstein cows were used in an experiment to evaluate the effect of supplementing diets with either a plant- or an animal-based source of rumen-undegradable protein (RUP), with or without AA supplementation, during the transition period and early lactation on milk production response. The experimental design was a randomized block design with approximately one-third of the cows being primiparous. Cows were assigned to 1 of 4 prepartum diets introduced 3 wk before the expected calving date and switched to the corresponding postpartum diet at calving. Diets 1 (AMI) and 2 (AMI+) included a vegetable RUP source (heat- and lignosulfonate-treated canola meal), with diet 2 containing supplemental Lys x HCl and Met hydroxy analog sources [D,L-2 hydroxy-4-(methylthio)-butanoic acid; Alimet feed supplement]. Diets 3 (PRO) and 4 (PRO+) consisted of a blend of animal RUP sources (blood meal, fish meal, feather meal, and porcine meat and bone meal), with diet 4 containing supplemental Lys x HCl and Met hydroxy analog sources [D,L-2 hydroxy-4-(methylthio)-butanoic acid; Alimet]. During the first 4 wk of lactation, dry matter intake was less when synthetic Lys x HCl and Alimet were supplemented, but this effect was no longer evident in wk 5 to 9 of the experiment. Interestingly, despite the initial decrease in dry matter intake in the cows fed AA-supplemented diets, there was no effect of treatment on milk production or the ratio of fat-corrected milk to dry matter intake throughout the 17 wk of the study. Undegradable protein source (vegetable vs. animal) did not affect dry matter intake, milk production, or 3.5% fat-corrected milk production for the first 17 wk of lactation. The results of this study indicate that heat- and lignosulfonate-treated canola meal can be used as a source of undegradable protein in place of high-quality rumen-undegradable animal protein sources without negative effects on milk production when diets are equivalent

  18. Conserved methionine dictates substrate preference in Nramp-family divalent metal transporters

    PubMed Central

    Bozzi, Aaron T.; Bane, Lukas B.; Weihofen, Wilhelm A.; McCabe, Anne L.; Singharoy, Abhishek; Chipot, Christophe J.; Schulten, Klaus

    2016-01-01

    Natural resistance-associated macrophage protein (Nramp) family transporters catalyze uptake of essential divalent transition metals like iron and manganese. To discriminate against abundant competitors, the Nramp metal-binding site should favor softer transition metals, which interact either covalently or ionically with coordinating molecules, over hard calcium and magnesium, which interact mainly ionically. The metal-binding site contains an unusual, but conserved, methionine, and its sulfur coordinates transition metal substrates, suggesting a vital role in their transport. Using a bacterial Nramp model system, we show that, surprisingly, this conserved methionine is dispensable for transport of the physiological manganese substrate and similar divalents iron and cobalt, with several small amino acid replacements still enabling robust uptake. Moreover, the methionine sulfur’s presence makes the toxic metal cadmium a preferred substrate. However, a methionine-to-alanine substitution enables transport of calcium and magnesium. Thus, the putative evolutionary pressure to maintain the Nramp metal-binding methionine likely exists because it—more effectively than any other amino acid—increases selectivity for low-abundance transition metal transport in the presence of high-abundance divalents like calcium and magnesium. PMID:27573840

  19. Parasites suppress immune-enhancing effect of methionine in nestling great tits.

    PubMed

    Wegmann, Michèle; Voegeli, Beatrice; Richner, Heinz

    2015-01-01

    After birth, an organism needs to invest both in somatic growth and in the development of efficient immune functions to counter the effects of pathogens, and hence an investment trade-off is predicted. To explore this trade-off, we simultaneously exposed nestling great tits (Parus major) to a common ectoparasite, while stimulating immune function. Using a 2 × 2 experimental design, we first infested half of the nests with hen fleas (Ceratophyllus gallinae) on day 3 post-hatch and later, on day 9-13 post-hatch, and then supplemented half of the nestlings within each nest with an immuno-enhancing amino acid (methionine). We then assessed the non-specific immune response by measuring both the inflammatory response to a lipopolysaccharide (LPS) and assessing the levels of acute phase proteins (APP). In parasite-infested nestlings, methionine had a negative effect on body mass close to fledging. Methionine had an immune-enhancing effect in the absence of ectoparasites only. The inflammatory response to LPS was significantly lower in nestlings infested with fleas and was also lower in nestlings supplemented with methionine. These patterns of immune responses suggest an immunosuppressive effect of ectoparasites that could neutralise the immune-enhancing effect of methionine. Our study thus suggests that the trade-off between investment in life history traits and immune function is only partly dependent on available resources, but shows that parasites may influence this trade-off in a more complex way, by also inhibiting important physiological functions.

  20. Methionine supplementation influences melanin-based plumage colouration in Eurasian kestrel, Falco tinnunculus, nestlings.

    PubMed

    Parejo, Deseada; Silva, Nadia

    2009-11-01

    The extent to which the expression of melanin-based plumage colouration in birds is genetically or environmentally determined is controversial. Here, we performed a between-nest design supplementation with either the sulphur amino acid dl-methionine or with water to investigate the importance of the non-genetic component of melanin-based plumage colouration in the Eurasian kestrel, Falco tinnunculus. Methionine affects growth and immunity, thus we aimed to modify nestling growth and immunity before feather development. Then, we measured the effect of the experiment on colouration of two melanin-based plumage patches of nestling kestrels. We found that methionine slowed down nestling growth through treatment administration and that nestlings compensated by speeding up their growth later. We did not find any effects of methionine on nestling immunity (i.e. lymphocyte counts, natural antibody levels or complement-mediated immunity). Effects on growth seemed to be mirrored by changes in nestling colouration in the two sexes: methionine-nestlings showed less intense brown plumage on their backs compared with control nestlings. These results provide support for a non-genetic determination of a melanin-based plumage patch in the two sexes of nestling kestrels.

  1. Amino acid metabolism and protein synthesis in malarial parasites*

    PubMed Central

    Sherman, I. W.

    1977-01-01

    Malaria-infected red cells and free parasites have limited capabilities for the biosynthesis of amino acids. Therefore, the principal amino acid sources for parasite protein synthesis are the plasma free amino acids and host cell haemoglobin. Infected cells and plasmodia incorporate exogenously supplied amino acids into protein. However, the hypothesis that amino acid utilization (from an external source) is related to availability of that amino acid in haemoglobin is without universal support: it is true for isoleucine and for Plasmodium knowlesi and P. falciparum, but not for methionine, cysteine, and other amino acids, and it does not apply to P. lophurae. More by default than by direct evidence, haemoglobin is believed to be the main amino acid reservoir available to the intraerythrocytic plasmodium. Haemoglobin, ingested via the cytostome, is held in food vacuoles where auto-oxidation takes place. As a consequence, haem is released and accumulates in the vacuole as particulate haemozoin (= malaria pigment). Current evidence favours the view that haemozoin is mainly haematin. Acid and alkaline proteases (identified in crude extracts from mammalian and avian malarias) are presumably secreted directly into the food vacuole. They then digest the denatured globin and the resulting amino acids are incorporated into parasite protein. Cell-free protein synthesizing systems have been developed using P. knowlesi and P. lophurae ribosomes. In the main these systems are typically eukaryotic. Studies of amino acid metabolism are exceedingly limited. Arginine, lysine, methionine, and proline are incorporated into protein, whereas glutamic acid is metabolized via an NADP-specific glutamic dehydrogenase. Glutamate oxidation generates NADPH and auxiliary energy (in the form of α-ketoglutarate). The role of red cell glutathione in the economy of the parasite remains obscure. Important goals for future research should be: quantitative assessment of the relative importance of

  2. Utilization of supplemental methionine sources by primary cultures of chick hepatocytes

    SciTech Connect

    Dibner, J.J.

    1983-10-01

    Utilization of 2-hydroxy-4-(methylthio) butanoic acid (HMB) as a substrate for protein synthesis was studied by using primary cultures of chick liver cells. Cultures were prepared by enzymatic dissociation of livers from week old Hubbard broiler chicks and were maintained for 4 days under nonproliferative conditions. Hepatocyte differentiation was verified by using dexamethasone induction of tyrosine aminotransferase activity. Conversion of (14C)HMB to L-methionine was shown by chromatographic analysis of hepatocyte protein hydrolysate and incorporation into protein was proven by cycloheximide inhibition of synthesis. When incorporation of HMB was compared to that of DL-methionine (DLM) equimolar quantities of the two sources were found in liver cell protein. These results support, at a cellular level, the conclusion that HMB and DLM are biochemically equivalent sources of methionine for protein synthesis.

  3. Fluoresence quenching of riboflavin in aqueous solution by methionin and cystein

    NASA Astrophysics Data System (ADS)

    Drössler, P.; Holzer, W.; Penzkofer, A.; Hegemann, P.

    2003-01-01

    The fluorescence quantum distributions, fluorescence quantum yields, and fluorescence lifetimes of riboflavin in methanol, DMSO, water, and aqueous solutions of the sulphur atom containing amino acids methionin and cystein have been determined. In methanol, DMSO, and water (pH=4-8) only dynamic fluorescence reduction due to intersystem crossing and internal conversion is observed. In aqueous methionin solutions of pH=5.25-9 a pH independent static and dynamic fluorescence quenching occurs probably due to riboflavin anion-methionin cation pair formation. In aqueous cystein solutions (pH range from 4.15 to 9) the fluorescence quenching increases with rising pH due to cystein thiolate formation. The cystein thiol form present at low pH does not react with neutral riboflavin. Cystein thiolate present at high pH seems to react with neutral riboflavin causing riboflavin deprotonation (anion formation) by cystein thiolate reduction to the cystein thiol form.

  4. Identification and characterization of a methionine γ-lyase in the calicheamicin biosynthetic cluster of Micromonospora echinospora.

    PubMed

    Song, Haigang; Xu, Ri; Guo, Zhihong

    2015-01-02

    CalE6 is a previously uncharacterized protein involved in the biosynthesis of calicheamicins in Micromonospora echinospora. It is a pyridoxal-5'-phosphate-dependent enzyme and exhibits high sequence homology to cystathionine γ-lyases and cystathionine γ-synthases. However, it was found to be active towards methionine and to convert this amino acid into α-ketobutyrate, ammonium, and methanethiol. The crystal structure of the cofactor-bound holoenzyme was resolved at 2.0 Å; it contains two active site residues, Gly105 and Val322, specific for methionine γ-lyases. Modeling of methionine into the active site allows identification of the active site residues responsible for substrate recognition and catalysis. These findings support that CalE6 is a putative methionine γ-lyase producing methanethiol as a building block in biosynthesis of calicheamicins.

  5. Amperometric biosensor based on diamond paste for the enantioanalysis of L-lysine.

    PubMed

    Stefan-van Staden, Raluca-Ioana; Nejem, R'afat Mahmoud; van Staden, Jacobus Frederick; Aboul-Enein, Hassan Y

    2012-05-15

    An amperometric biosensor was proposed for the enantioanalysis of L-lysine. The biosensor is based on the impregnation of L-lysine oxidase in diamond paste. The potential used for the determination of l-lysine was 650 mV. The biosensor exhibited a linear concentration range between 1 and 100 nmol/L with a limit of detection of 4 pmol/L. The selectivity of the biosensor is high over other amino acids, such as L-serine, L-leucine, L-aspartic acid, L-glutamic acid, histamine, glycine. The proposed biosensor can be applied for the determination of L-lysine in serum samples and pharmaceutical compounds.

  6. Cloning and sequence analysis of a cDNA encoding a Brazil nut protein exceptionally rich in methionine.

    PubMed

    Altenbach, S B; Pearson, K W; Leung, F W; Sun, S S

    1987-05-01

    The primary amino acid sequence of an abundant methionine-rich seed protein found in Brazil nut (Bertholletia excelsa H.B.K.) has been elucidated by protein sequencing and from the nucleotide sequence of cDNA clones. The 9 kDa subunit of this protein was found to contain 77 amino acids of which 14 were methionine (18%) and 6 were cysteine (8%). Over half of the methionine residues in this subunit are clustered in two regions of the polypeptide where they are interspersed with arginine residues. In one of these regions, methionine residues account for 5 out of 6 amino acids and four of these methionine residues are contiguous. The sequence data verifies that the Brazil nut sulfur-rich protein is synthesized as a precursor polypeptide that is considerably larger than either of the two subunits of the mature protein. Three proteolytic processing steps by which the encoded polypeptide is sequentially trimmed to the 9 kDa and 3 kDa subunit polypeptides have been correlated with the sequence information. In addition, we have found that the sulfur-rich protein from Brazil nut is homologous in its amino acid sequence to small water-soluble proteins found in two other oilseeds, castor bean (Ricinus communis) and rapeseed (Brassica napus). When the amino acid sequences of these three proteins are aligned to maximize homology, the arrangement of cysteine residues is conserved. However, the two subunits of the Brazil nut protein contain over 19% methionine whereas the homologous proteins from castor bean and rapeseed contain only 2.1% and 2.6% methionine, respectively.

  7. Lysine malonylation is elevated in type 2 diabetic mouse models and enriched in metabolic associated proteins.

    PubMed

    Du, Yipeng; Cai, Tanxi; Li, Tingting; Xue, Peng; Zhou, Bo; He, Xiaolong; Wei, Peng; Liu, Pingsheng; Yang, Fuquan; Wei, Taotao

    2015-01-01

    Protein lysine malonylation, a newly identified protein post-translational modification (PTM), has been proved to be evolutionarily conserved and is present in both eukaryotic and prokaryotic cells. However, its potential roles associated with human diseases remain largely unknown. In the present study, we observed an elevated lysine malonylation in a screening of seven lysine acylations in liver tissues of db/db mice, which is a typical model of type 2 diabetes. We also detected an elevated lysine malonylation in ob/ob mice, which is another model of type 2 diabetes. We then performed affinity enrichment coupled with proteomic analysis on liver tissues of both wild-type (wt) and db/db mice and identified a total of 573 malonylated lysine sites from 268 proteins. There were more malonylated lysine sites and proteins in db/db than in wt mice. Five proteins with elevated malonylation were verified by immunoprecipitation coupled with Western blot analysis. Bioinformatic analysis of the proteomic results revealed the enrichment of malonylated proteins in metabolic pathways, especially those involved in glucose and fatty acid metabolism. In addition, the biological role of lysine malonylation was validated in an enzyme of the glycolysis pathway. Together, our findings support a potential role of protein lysine malonylation in type 2 diabetes with possible implications for its therapy in the future.

  8. Lysine Malonylation Is Elevated in Type 2 Diabetic Mouse Models and Enriched in Metabolic Associated Proteins*

    PubMed Central

    Du, Yipeng; Cai, Tanxi; Li, Tingting; Xue, Peng; Zhou, Bo; He, Xiaolong; Wei, Peng; Liu, Pingsheng; Yang, Fuquan; Wei, Taotao

    2015-01-01

    Protein lysine malonylation, a newly identified protein post-translational modification (PTM), has been proved to be evolutionarily conserved and is present in both eukaryotic and prokaryotic cells. However, its potential roles associated with human diseases remain largely unknown. In the present study, we observed an elevated lysine malonylation in a screening of seven lysine acylations in liver tissues of db/db mice, which is a typical model of type 2 diabetes. We also detected an elevated lysine malonylation in ob/ob mice, which is another model of type 2 diabetes. We then performed affinity enrichment coupled with proteomic analysis on liver tissues of both wild-type (wt) and db/db mice and identified a total of 573 malonylated lysine sites from 268 proteins. There were more malonylated lysine sites and proteins in db/db than in wt mice. Five proteins with elevated malonylation were verified by immunoprecipitation coupled with Western blot analysis. Bioinformatic analysis of the proteomic results revealed the enrichment of malonylated proteins in metabolic pathways, especially those involved in glucose and fatty acid metabolism. In addition, the biological role of lysine malonylation was validated in an enzyme of the glycolysis pathway. Together, our findings support a potential role of protein lysine malonylation in type 2 diabetes with possible implications for its therapy in the future. PMID:25418362

  9. Lysine-overproducing mutants of Saccharomyces cerevisiae baker's yeast isolated in continuous culture.

    PubMed Central

    Gasent-Ramírez, J M; Benítez, T

    1997-01-01

    Saccharomyces cerevisiae baker's yeast mutants which produce 3 to 17 times as much lysine as the wild type, depending on the nitrogen source, have been selected. The baker's yeast strain was growth in a pH-regulated chemostat in minimal medium with proline as the nitrogen source, supplemented with increasing concentrations of the toxic analog of the lysine S-2-aminoethyl-L-cysteine (AEC). The lysine-overproducing mutants, which were isolated as AEC-resistant mutants, were also resistant to high external concentrations of lysine and to alpha-aminoadipate and seemed to be affected in the lysine biosynthetic pathway but not in the biosynthetic pathways of other amino acids. Lysine overproduction by one of the mutants seemed to be due to, at least, the loss of repression of the homocitrate synthase encoded by the LYS20 gene. The mutant grew slower than the wild type, and its dough-raising capacity was reduced in in vitro assays, probably due to the toxic effects of lysine accumulation or of an intermediate produced in the pathway. This mutant can be added as a food supplement to enrich the nutritive qualities of bakery products, and its resistance to alpha-aminoadipate, AEC, and lysine can be used as a dominant marker. PMID:9406398

  10. Antioxidant activity of amino acids in soybean oil at frying temperature: Structural effects and synergism with tocopherols.

    PubMed

    Hwang, Hong-Sik; Winkler-Moser, Jill K

    2017-04-15

    The purpose of this study was to evaluate amino acids as natural antioxidants for frying. Twenty amino acids were added to soybean oil heated to 180°C, and the effects of amino acid structure on the antioxidant activity were investigated. Amino acids containing a thiol, a thioether, or an extra amine group such as arginine, cysteine, lysine, methionine, and tryptophan had the strongest antioxidant activities. At 5.5mM, these amino acids had stronger antioxidant activities than 0.02% (1.1mM) tert-butylhydroquinone (TBHQ). A functional group such as an amide, carboxylic acid, imidazole, or phenol appeared to negatively affect amino acid antioxidant activity. Synergism between amino acids and tocopherols was demonstrated, and we found that this synergistic interaction may be mostly responsible for the antioxidant activity that was observed. In a frying study with potato cubes, 5.5mM l-methionine had significantly stronger antioxidant activity than 0.02% TBHQ.

  11. 21 CFR 172.399 - Zinc methionine sulfate.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... 21 Food and Drugs 3 2013-04-01 2013-04-01 false Zinc methionine sulfate. 172.399 Section 172.399... CONSUMPTION Special Dietary and Nutritional Additives § 172.399 Zinc methionine sulfate. Zinc methionine... conditions: (a) The additive is the product of the reaction between equimolar amounts of zinc sulfate and...

  12. 21 CFR 582.5475 - Methionine.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... 21 Food and Drugs 6 2012-04-01 2012-04-01 false Methionine. 582.5475 Section 582.5475 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) ANIMAL DRUGS, FEEDS, AND RELATED PRODUCTS SUBSTANCES GENERALLY RECOGNIZED AS SAFE Nutrients and/or Dietary...

  13. 21 CFR 582.5475 - Methionine.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... 21 Food and Drugs 6 2011-04-01 2011-04-01 false Methionine. 582.5475 Section 582.5475 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) ANIMAL DRUGS, FEEDS, AND RELATED PRODUCTS SUBSTANCES GENERALLY RECOGNIZED AS SAFE Nutrients and/or Dietary...

  14. 21 CFR 582.5475 - Methionine.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... 21 Food and Drugs 6 2014-04-01 2014-04-01 false Methionine. 582.5475 Section 582.5475 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) ANIMAL DRUGS, FEEDS, AND RELATED PRODUCTS SUBSTANCES GENERALLY RECOGNIZED AS SAFE Nutrients and/or Dietary...

  15. 21 CFR 582.5475 - Methionine.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... 21 Food and Drugs 6 2013-04-01 2013-04-01 false Methionine. 582.5475 Section 582.5475 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) ANIMAL DRUGS, FEEDS, AND RELATED PRODUCTS SUBSTANCES GENERALLY RECOGNIZED AS SAFE Nutrients and/or Dietary...

  16. 21 CFR 582.5475 - Methionine.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 21 Food and Drugs 6 2010-04-01 2010-04-01 false Methionine. 582.5475 Section 582.5475 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) ANIMAL DRUGS, FEEDS, AND RELATED PRODUCTS SUBSTANCES GENERALLY RECOGNIZED AS SAFE Nutrients and/or Dietary...

  17. The development and amino acid binding ability of nano-materials based on azo derivatives: theory and experiment.

    PubMed

    Shang, Xuefang; Du, Jinge; Yang, Wancai; Liu, Yun; Fu, Zhiyuan; Wei, Xiaofang; Yan, Ruifang; Yao, Ningcong; Guo, Yaping; Zhang, Jinlian; Xu, Xiufang

    2014-05-01

    Two nano-material-containing azo groups have been designed and developed, and the binding ability of nano-materials with various amino acids has been characterized by UV-vis and fluorescence titrations. Results indicated that two nano-materials showed the strongest binding ability for homocysteine among twenty normal kinds of amino acids (alanine, valine, leucine, isoleucine, methionine, aspartic acid, glutamic acid, arginine, glycine, serine, threonine, asparagine, phenylalanine, histidine, tryptophan, proline, lysine, glutamine, tyrosine and homocysteine). The reason for the high sensitivity for homocysteine was that two nano-materials containing an aldehyde group reacted with SH in homocysteine and afforded very stable thiazolidine derivatives. Theoretical investigation further illustrated the possible binding mode in host-guest interaction and the roles of molecular frontier orbitals in molecular interplay. Thus, the two nano-materials can be used as optical sensors for the detection of homocysteine.

  18. Lysine fortification reduces anxiety and lessens stress in family members in economically weak communities in Northwest Syria.

    PubMed

    Smriga, Miro; Ghosh, Shibani; Mouneimne, Youssef; Pellett, Peter L; Scrimshaw, Nevin S

    2004-06-01

    Lysine is a limiting amino acid in diets based on wheat as the staple. In experimental animals, prolonged dietary lysine inadequacy increases stress-induced anxiety. If observed in humans, such a result would have a strong implication for the relationship between nutrition and communal quality of life and mental health. As part of a 3-month randomized double-blind study, we tested whether lysine fortification of wheat reduces anxiety and stress response in family members in poor Syrian communities consuming wheat as a staple food. In the lysine-fortified group, the plasma cortisol response to the blood drawing as a cause of stress was reduced in females, as was sympathetic arousal in males as measured by skin conductance. Lysine fortification also significantly reduced chronic anxiety as measured by the trait anxiety inventory in males. These results suggest that some stress responses in economically weak populations consuming cereal-based diets can be improved with lysine fortification.

  19. Identification, expression and antibacterial activities of an antimicrobial peptide NK-lysin from a marine fish Larimichthys crocea.

    PubMed

    Zhou, Qi-Jia; Wang, Jun; Liu, Min; Qiao, Ying; Hong, Wan-Shu; Su, Yong-Quan; Han, Kun-Huang; Ke, Qiao-Zhen; Zheng, Wei-Qiang

    2016-08-01

    As fundamental immunologic mechanism, the innate immunity system is more important than the specific immunity system in teleost fishes during pathogens infection. Antimicrobial peptides are integral parts of the innate immune system, and play significant roles against pathogens infection. NK-lysin, the compounds of the natural killer cells and cytotoxic T cells, are potent and effective antimicrobial peptides widely distributed in animals. In this study, we reported the sequence characteristics, expression profiles and antibacterial activities of a NK-lysin gene (Lc-NK-lysin) from a commercially important marine fish, the large yellow croaker (Larimichthys crocea). The open reading frame of Lc-NK-lysin cDNA sequence was 447 bp in length, coding 148 amino acids. The genomic DNA of Lc-NK-lysin has the common features of NK-lysin family, consisting of five exons and four introns, and in its deduced mature peptide, there are six well-conserved cysteine residues and a Saposin B domain. Lc-NK-lysin was expressed in all tested tissues (skin, muscle, gill, brain, head kidney, heart, liver, spleen, stomach and intestine) with different expression patterns. In pathogens infection the expression profiles of Lc-NK-lysin varied significantly in gill, head kidney, spleen and liver, indicating its role in immune response. Two peptides (Lc-NK-lysin-1 and Lc-NK-lysin-2) divided from the core region of the Lc-NK-lysin mature polypeptide were chemically synthesized and their antibacterial activities were examined; the potential function on the inhibition of bacteria propagation was revealed. Our results suggested that Lc-NK-lysin is a typical member of the NK-lysin family and as an immune-related gene it involves in the immune response when pathogens invasion.

  20. NOVEL ANTI-MICROBIAL PEPTIDE, NK-LYSIN, IS PRODUCED LOCALLY IN THE GUT OF EIMERIA-INFECTED HOST

    Technology Transfer Automated Retrieval System (TEKTRAN)

    NK-lysin is an anti-microbial and anti-tumor protein produced by NK cells and T lymphocytes in mammals and is considered to be an important component of the local innate immune response to pathogens. Chicken NK-lysin consists of an 868 bp DNA sequence with an ORF of 140 amino acids with a predicted ...

  1. Coacervate-like microspheres from lysine-rich proteinoid

    NASA Technical Reports Server (NTRS)

    Rohlfing, D. L.

    1975-01-01

    Microspheres form isothermally from lysine-rich proteinoid when the ionic strength of the solution is increased with NaCl or other salts. Studies with different monovalent anions and with polymers of different amino acid composition indicate that charge neutralization and hydrophobic bonding contribute to microsphere formation. The particles also form in sea water, especially if heated or made slightly alkaline. The microspheres differ from those made from acidic proteinoid but resemble coacervate droplets in some ways (isothermal formation, limited stability, stabilization by quinone, uptake of dyes). Because the constituent lysine-rich proteinoid is of simulated prebiotic origin, the study is interpreted to add emphasis to and suggest an evolutionary continuity for coacervation phenomena.

  2. Cobalamin inactivation by nitrous oxide produces severe neurological impairment in fruit bats: protection by methionine and aggravation by folates

    SciTech Connect

    van der Westhuyzen, J.; Fernandes-Costa, F.; Metz, J.

    1982-11-01

    Nitrous oxide, which inactivates cobalamin when administered to fruit bats, results in severe neurological impairment leading to ataxia, paralysis and death. This occurs after about 6 weeks in animals depleted of cobalamin by dietary restriction, and after about 10 weeks in cobalamin replete bats. Supplementation of the diet with pteroylglutamic acid caused acceleration of the neurological impairment--the first unequivocal demonstration of aggravation of the neurological lesion in cobalamin deficiency by pteroylglutamic acid. The administration of formyltetrahydropteroylglutamic acid produced similar aggravation of the neurological lesion. Supplementation of the diet with methionine protected the bats from neurological impairment, but failed to prevent death. Methionine supplementation protected against the exacerbating effect of folate, preventing the development of neurological changes. These findings lend support to the hypothesis that the neurological lesion in cobalamin deficiency may be related to a deficiency in the methyl donor S-adenosylmethionine which follows diminished synthesis of methionine.

  3. Studies on the biochemical composition, energetics and essential amino acids of three mudskippers in Xiamen Harbour

    NASA Astrophysics Data System (ADS)

    Jun, Wang; Yong-Quan, Su

    1995-12-01

    Measurements were made on the contents of protein, lipid, glycogen (PLG) and water, and on caloric values and amino acids, in muscle of three mudskippers Periophthalmus cantonensis, Scarteiaos viridis and Boleophthalmus pectinirostris collected from Haicang, Xiamen. The essential amino acids (EAA) for these fishes were also studied with radioisotopic trace method. The results showed: (1) The content of each component in tested fish muscles differed slightly, and protein was the most important component making up from 6.685% to 9.891% of the wet weight (about 44.21%-50.45% of dry weight); (2) Energy calculated from the sum of protein, lipid and glycogen in wet muscle was low (<4.3kJ/g) in these fishes, especially in B. pectinirostris (<3.1 kJ/g); the ratios of energy to protein content ( E/P) also were low (<39.873-45.535kJ/g); (3) Seventeen amino acids were determined in these three fishes. The content of the same amino acid (among the seventeen) tested in different species and sexes varied slightly. The amounts of methionine, phenylalanine lysine, arginine, histidine, threonine isoleucine and leucine which are indispensable for the needs of human beings and animals were relatively high, accounting for 47.35%-48.06% of the total amino acid content. (4) Leucine, isoleucine, arginine, lysine, phenylalanine, tryptophan, methionine, histidine, threonine, and valine, are essential in the diet of the three mudskippers as the radioisotopic trace method using D-[U-14C]-glucose showed little or no radioactivity was incorporated into these ten amino acids.

  4. Studies on the biochemical composition, energetics and essential amino acids of three mudskippers in Xiamen Harbour

    NASA Astrophysics Data System (ADS)

    Wang, Jun; Su, Yong-Quan

    1995-12-01

    Measurements were made on the contents of protein, lipid, glycogen (PLG) and water, and on caloric values and amino acids, in muscle of three mudskippers Periophthalmus cantonensis, Scarteiaos viridis and Boleophthalmus pectinirostris collected from Haicang, Xiamen. The essential amino acids (EAA) for these fishes were also studied with radioisotopic trace method. The results showed: (1) The content of each component in tested fish muscles differed slightly, and protein was the most important component making up from 6.685% to 9.891% of the wet weight (about 44.21% 50.45% of dry weight); (2) Energy calculated from the sum of protein, lipid and glycogen in wet muscle was low (<4.3kJ/g) in these fishes, especially in B. pectinirostris (<3.1 kJ/g); the ratios of energy to protein content ( E/P) also were low (<39.873 45.535kJ/g); (3) Seventeen amino acids were determined in these three fishes. The content of the same amino acid (among the seventeen) tested in different species and sexes varied slightly. The amounts of methionine, phenylalanine lysine, arginine, histidine, threonine isoleucine and leucine which are indispensable for the needs of human beings and animals were relatively high, accounting for 47.35% 48.06% of the total amino acid content. (4) Leucine, isoleucine, arginine, lysine, phenylalanine, tryptophan, methionine, histidine, threonine, and valine, are essential in the diet of the three mudskippers as the radioisotopic trace method using D-[U-14C]-glucose showed little or no radioactivity was incorporated into these ten amino acids.

  5. Nitrogen retention and performance of brown laying hens on diets with different protein content and constant concentration of amino acids and energy.

    PubMed

    Meluzzi, A; Sirri, F; Tallarico, N; Franchini, A

    2001-05-01

    1. The aim of this study was to determine the nitrogen balance and the performance of laying hens fed on diets with a protein content lower than the diets currently used in commercial practice but with adequate concentrations of lysine, sulphur amino acids, tryptophan and threonine. 2. Ninety-six Hy-Line Brown hens, 24 weeks old, were divided into 3 groups of 8 replicates and received, for 16 weeks, diets formulated to have 3 different protein concentrations: 170 (control), 150 and 130 g/kg CP and the same energy content. For each protein concentration, the contents of lysine, methionine, methionine+cystine, tryptophan and threonine were maintained at minimum requirement concentrations by supplying synthetic amino acids. 3. In the first half of the trial, egg production and egg weight were similar in all groups. From the 9th week onwards group 150 CP laid heavier eggs and had a slightly lower egg deposition and total mass. Food conversion ratio was best in the control group. 4. Nitrogen intake was related to the protein concentration of the diet, the food intake being almost the same in the 3 experimental groups. Faecal nitrogen content significantly and linearly decreased with reduction in dietary protein content and was about 50% of the intake. Considering the nitrogen faecal/intake ratio, the 150 CP group showed better nitrogen utilisation at each sampling time.

  6. Global Proteomics Analysis of Protein Lysine Methylation

    PubMed Central

    Cao, Xing-Jun; Garcia, Benjamin A.

    2017-01-01

    Lysine methylation is a common protein post-translational modification dynamically mediated by protein lysine methyltransferases (PKMTs) and demethylases (PKDMs). Beyond histone proteins, lysine methylation on non-histone proteins play substantial roles in a variety of functions in cells, and is closely associated with diseases such as cancer. A large body of evidence indicates that the dysregulation of some PKMTs lead to tumorigenesis via their non-histone substrates. However, more studies on other PKMTs have made slow progress owing to the lack of the approaches for extensive screening of lysine methylation sites. Recently a series of publications to perform large-scale analysis of protein lysine methylation have emerged. In this unit, we introduce a protocol for the global analysis of protein lysine methylation in cells by means of immunoaffinity enrichment and mass spectrometry. PMID:27801517

  7. A Methionine Deficient Diet Enhances Adipose Tissue Lipid Metabolism and Alters Anti-Oxidant Pathways in Young Growing Pigs

    PubMed Central

    Castellano, Rosa; Perruchot, Marie-Hélène; Conde-Aguilera, José Alberto; van Milgen, Jaap; Collin, Anne; Tesseraud, Sophie; Mercier, Yves; Gondret, Florence

    2015-01-01

    Methionine is a rate-limiting amino-acid for protein synthesis but non-proteinogenic roles on lipid metabolism and oxidative stress have been demonstrated. Contrary to rodents where a dietary methionine deficiency led to a lower adiposity, an increased lipid accretion rate has been reported in growing pigs fed a methionine deficient diet. This study aimed to clarify the effects of a dietary methionine deficiency on different aspects of tissue lipid metabolism and anti-oxidant pathways in young pigs. Post-weaned pigs (9.8 kg initial body weight) were restrictively-fed diets providing either an adequate (CTRL) or a deficient methionine supply (MD) during 10 days (n=6 per group). At the end of the feeding trial, pigs fed the MD diet had higher lipid content in subcutaneous adipose tissue. Expression levels of genes involved in glucose uptake, lipogenesis but also lipolysis, and activities of NADPH enzyme suppliers were generally higher in subcutaneous and perirenal adipose tissues of MD pigs, suggesting an increased lipid turnover in those pigs. Activities of the anti-oxidant enzymes superoxide dismutase, catalase and glutathione reductase were increased in adipose tissues and muscle of MD pigs. Expression level and activity of the glutathione peroxidase were also higher in liver of MD pigs, but hepatic contents in the reduced and oxidized forms of glutathione and glutathione reductase activity were lower compared with control pigs. In plasma, superoxide dismutase activity was higher but total anti-oxidant power was lower in MD pigs. These results show that a dietary methionine deficiency resulted in increased levels of lipogenesis and lipolytic indicators in porcine adipose tissues. Decreased glutathione content in the liver and coordinated increase of enzymatic antioxidant activities in adipose tissues altered the cellular redox status of young pigs fed a methionine-deficient diet. These findings illustrate that a rapidly growing animal differently adapts tissue

  8. Changes in plasma amino acid concentrations with increasing age in patients with propionic acidemia.

    PubMed

    Scholl-Bürgi, Sabine; Sass, Jörn Oliver; Heinz-Erian, Peter; Amann, Edda; Haberlandt, Edda; Albrecht, Ursula; Ertl, Claudia; Sigl, Sara Baumgartner; Lagler, Florian; Rostasy, Kevin; Karall, Daniela

    2010-05-01

    The objective of the study is to analyze plasma amino acid concentrations in propionic acidemia (PA) for the purpose of elucidating possible correlations between propionyl-CoA carboxylase deficiency and distinct amino acid behavior. Plasma concentrations of 19 amino acids were measured in 240 random samples from 11 patients (6 families) with enzymatically and/or genetically proven propionic acidemia (sampling period, January 2001-December 2007). They were compared with reference values from the literature and correlated with age using the Pearson correlation coefficient test. Decreased plasma concentrations were observed for glutamine, histidine, threonine, valine, isoleucine, leucine, phenylalanine and arginine. Levels of glycine, alanine and aspartate were elevated, while values of serine, asparagine, ornithine and glutamate were normal. For lysine, proline and methionine a clear association was not possible. Significant correlations with age were observed for 13 amino acids (positive correlation: asparagine, glutamine, proline, alanine, histidine, threonine, methionine, arginine; negative correlation: leucine, phenylalanine, ornithine, glutamate and aspartate). This study gives new insight over long-term changes in plasma amino acid concentrations and may provide options for future therapies (e.g., substitution of anaplerotic substances) in PA patients.

  9. Separation and detection of amino acid metabolites of Escherichia coli in microbial fuel cell with CE.

    PubMed

    Wang, Wei; Ma, Lihong; Lin, Ping; Xu, Kaixuan

    2016-07-01

    In this work, CE-LIF was employed to investigate the amino acid metabolites produced by Escherichia coli (E. coli) in microbial fuel cell (MFC). Two peptides, l-carnosine and l-alanyl-glycine, together with six amino acids, cystine, alanine, lysine, methionine, tyrosine, arginine were separated and detected in advance by a CE-LIF system coupled with a homemade spontaneous injection device. The injection device was devised to alleviate the effect of electrical discrimination for analytes during sample injection. All analytes could be completely separated within 8 min with detection limits of 20-300 nmol/L. Then this method was applied to analyze the substrate solution containing amino acid metabolites produced by E. coli. l-carnosine, l-alanyl-glycine, and cystine were used as the carbon, nitrogen, and sulfur source for the E. coli culture in the MFC to investigate the amino acid metabolites during metabolism. Two MFCs were used to compare the activity of metabolism of the bacteria. In the sample collected at the running time 200 h of MFC, the amino acid methionine was discovered as the metabolite with the concentrations 23.3 μg/L.

  10. Dietary crude protein has minimal effect on the activity of selected enzymes of methionine catabolism in kittens fed diets near-limiting in methionine.

    PubMed

    Strieker, M J; Morris, J G; Avery, E H; Freedland, R A; Rogers, Q R

    2008-04-01

    Previous experiments have shown that increasing the dietary crude protein (CP) of cats does not increase urea cycle enzymes or alanine amino transferase as occurs in rats. Also when an essential amino acid (EAA) is limiting in a diet for growing kittens, the kittens do not exhibit an amino acid imbalance when other EAAs are added to the diet. To study the metabolic basis for these observations which are different from that found in omnivores and herbivores, the hypothesis that increased dietary CP decreases methionine catabolism, so more is spared for growth, was tested. Fifteen male kittens were randomly assigned to one of three dietary treatments. Each diet contained 2.5 g l-methionine/kg diet and 200, 300 or 500 g CP/kg diet. The livers and kidneys were removed and assayed for methionine transaminase (MTA), cystathionase (CASE) and cystathionine synthase (CS). Free amino acid concentrations were determined in liver, kidney and plasma. The 300 and 500 g CP/kg groups had significantly greater kidney weights and body weight gains than the 200 g CP/kg group. Hepatic MTA activity was lower in the 300 than the 200 or 500 g CP/kg groups (p < 0.05). Renal MTA and CASE activities were 35% and 50% greater, respectively, for the 500 g CP/kg group than for the 200 g CP/kg diet group (p < 0.05). Renal CS activities for the 300 and 500 g CP/kg groups were 29% (p > 0.05) and 38% (p < 0.05) greater, respectively, than the 200 g CP/kg group. Cyst(e)ine concentrations were lower in the livers of the 500 g CP/kg group than the 200 g CP/kg group (p < 0.05). Cystathionine was lower in plasma and kidney from the 500 g CP/kg diet group than from the 200 g CP/kg diet group (p < 0.05). It was concluded that the metabolic basis for the increased growth of kittens fed diets marginally limiting in methionine, with increasing concentrations of dietary CP, was not mediated through decreased enzyme activity associated with the catabolism of methionine, but was the result of an increase in

  11. Effect of inhibitor compounds on Nε-(carboxymethyl)lysine (CML) and Nε-(carboxyethyl)lysine (CEL) formation in model foods.

    PubMed

    Srey, Chou; Hull, George L J; Connolly, Lisa; Elliott, Christopher T; del Castillo, M Dolores; Ames, Jennifer M

    2010-11-24

    The possible adverse effects on health of diet-derived advanced glycation endproducts (AGEs) and advanced lipoxidation endproducts (ALEs) is of current interest. This study had the objective of determining the effects of the addition of AGE/ALE inhibitors and different types of sugar and cooking oil on Nε-(carboxymethyl)lysine (CML) and Nε-(carboxyethyl)lysine (CEL) formation in model foods (sponge cakes). The cake baked using glucose produced the highest level of CML (2.07±0.24 mmol/mol lysine), whereas the cake baked using fructose produced the highest concentration of CEL (25.1±0.15 mmol/mol lysine). There were no significant differences between CML concentrations formed in the cakes prepared using different types of cooking oil, but significant differences (P<0.001) were observed between the cakes prepared using different proportions of cooking oil. The cakes containing oil generated greater concentrations of CML than sucrose. α-Tocopherol and rutin did not inhibit CML and CEL formation. In contrast, ferulic acid and thiamin, thiamin monophosphate, and thiamin pyrophosphate reduced CML and CEL formation.

  12. Adding a Lysine Mimic in the Design of Potent Inhibitors of Histone Lysine Methyltransferases

    SciTech Connect

    Chang, Yanqi; Ganesh, Thota; Horton, John R.; Spannhoff, Astrid; Liu, Jin; Sun, Aiming; Zhang, Xing; Bedford, Mark T.; Shinkai, Yoichi; Snyder, James P.; Cheng, Xiaodong

    2010-07-19

    Dynamic histone lysine methylation involves the activities of modifying enzymes (writers), enzymes removing modifications (erasers), and readers of the histone code. One common feature of these activities is the recognition of lysines in methylated and unmethylated states, whether they are substrates, reaction products, or binding partners. We applied the concept of adding a lysine mimic to an established inhibitor (BIX-01294) of histone H3 lysine 9 methyltransferases G9a and G9a-like protein by including a 5-aminopentyloxy moiety, which is inserted into the target lysine-binding channel and becomes methylated by G9a-like protein, albeit slowly. The compound enhances its potency in vitro and reduces cell toxicity in vivo. We suggest that adding a lysine or methyl-lysine mimic should be considered in the design of small-molecule inhibitors for other methyl-lysine writers, erasers, and readers.

  13. Identification and characterization of lysine-methylated sites on histones and non-histone proteins.

    PubMed

    Lee, Tzong-Yi; Chang, Cheng-Wei; Lu, Cheng-Tzung; Cheng, Tzu-Hsiu; Chang, Tzu-Hao

    2014-06-01

    Protein methylation is a kind of post-translational modification (PTM), and typically takes place on lysine and arginine amino acid residues. Protein methylation is involved in many important biological processes, and most recent studies focused on lysine methylation of histones due to its critical roles in regulating transcriptional repression and activation. Histones possess highly conserved sequences and are homologous in most species. However, there is much less sequence conservation among non-histone proteins. Therefore, mechanisms for identifying lysine-methylated sites may greatly differ between histones and non-histone proteins. Nevertheless, this point of view was not considered in previous studies. Here we constructed two support vector machine (SVM) models by using lysine-methylated data from histones and non-histone proteins for predictions of lysine-methylated sites. Numerous features, such as the amino acid composition (AAC) and accessible surface area (ASA), were used in the SVM models, and the predictive performance was evaluated using five-fold cross-validations. For histones, the predictive sensitivity was 85.62% and specificity was 80.32%. For non-histone proteins, the predictive sensitivity was 69.1% and specificity was 88.72%. Results showed that our model significantly improved the predictive accuracy of histones compared to previous approaches. In addition, features of the flanking region of lysine-methylated sites on histones and non-histone proteins were also characterized and are discussed. A gene ontology functional analysis of lysine-methylated proteins and correlations of lysine-methylated sites with other PTMs in histones were also analyzed in detail. Finally, a web server, MethyK, was constructed to identify lysine-methylated sites. MethK now is available at http://csb.cse.yzu.edu.tw/MethK/.

  14. Increased methionine sulfoxide content of apoA-I in type 1 diabetes.

    PubMed

    Brock, Jonathan W C; Jenkins, Alicia J; Lyons, Timothy J; Klein, Richard L; Yim, Eunsil; Lopes-Virella, Maria; Carter, Rickey E; Thorpe, Suzanne R; Baynes, John W

    2008-04-01

    Cardiovascular disease is a major cause of morbidity and premature mortality in diabetes. HDL plays an important role in limiting vascular damage by removing cholesterol and cholesteryl ester hydroperoxides from oxidized low density lipoprotein and foam cells. Methionine (Met) residues in apolipoprotein A-I (apoA-I), the major apolipoprotein of HDL, reduce peroxides in HDL lipids, forming methionine sulfoxide [Met(O)]. We examined the extent and sites of Met(O) formation in apoA-I of HDL isolated from plasma of healthy control and type 1 diabetic subjects to assess apoA-I exposure to lipid peroxides and the status of oxidative stress in the vascular compartment in diabetes. Three tryptic peptides of apoA-I contain Met residues: Q(84)-M(86)-K(88), W(108)-M(112)-R(116), and L(144)-M(148)-R(149). These peptides and their Met(O) analogs were identified and quantified by mass spectrometry. Relative to controls, Met(O) formation was significantly increased at all three locations (Met(86), Met(112), and Met(148)) in diabetic patients. The increase in Met(O) in the diabetic group did not correlate with other biomarkers of oxidative stress, such as N(epsilon)-malondialdehyde-lysine or N(epsilon)-(carboxymethyl)lysine, in plasma or lipoproteins. The higher Met(O) content in apoA-I from diabetic patients is consistent with increased levels of lipid peroxidation products in plasma in diabetes. Using the methods developed here, future studies can address the relationship between Met(O) in apoA-I and the risk, development, or progression of the vascular complications of diabetes.

  15. Druggability of methyl-lysine binding sites

    NASA Astrophysics Data System (ADS)

    Santiago, C.; Nguyen, K.; Schapira, M.

    2011-12-01

    Structural modules that specifically recognize—or read—methylated or acetylated lysine residues on histone peptides are important components of chromatin-mediated signaling and epigenetic regulation of gene expression. Deregulation of epigenetic mechanisms is associated with disease conditions, and antagonists of acetyl-lysine binding bromodomains are efficacious in animal models of cancer and inflammation, but little is known regarding the druggability of methyl-lysine binding modules. We conducted a systematic structural analysis of readers of methyl marks and derived a predictive druggability landscape of methyl-lysine binding modules. We show that these target classes are generally less druggable than bromodomains, but that some proteins stand as notable exceptions.

  16. CPLM: a database of protein lysine modifications

    PubMed Central

    Liu, Zexian; Wang, Yongbo; Gao, Tianshun; Pan, Zhicheng; Cheng, Han; Yang, Qing; Cheng, Zhongyi; Guo, Anyuan; Ren, Jian; Xue, Yu

    2014-01-01

    We reported an integrated database of Compendium of Protein Lysine Modifications (CPLM; http://cplm.biocuckoo.org) for protein lysine modifications (PLMs), which occur at active ε-amino groups of specific lysine residues in proteins and are critical for orchestrating various biological processes. The CPLM database was updated from our previously developed database of Compendium of Protein Lysine Acetylation (CPLA), which contained 7151 lysine acetylation sites in 3311 proteins. Here, we manually collected experimentally identified substrates and sites for 12 types of PLMs, including acetylation, ubiquitination, sumoylation, methylation, butyrylation, crotonylation, glycation, malonylation, phosphoglycerylation, propionylation, succinylation and pupylation. In total, the CPLM database contained 203 972 modification events on 189 919 modified lysines in 45 748 proteins for 122 species. With the dataset, we totally identified 76 types of co-occurrences of various PLMs on the same lysine residues, and the most abundant PLM crosstalk is between acetylation and ubiquitination. Up to 53.5% of acetylation and 33.1% of ubiquitination events co-occur at 10 746 lysine sites. Thus, the various PLM crosstalks suggested that a considerable proportion of lysines were competitively and dynamically regulated in a complicated manner. Taken together, the CPLM database can serve as a useful resource for further research of PLMs. PMID:24214993

  17. Global Analysis of Lysine Acetylation Suggests the Involvement of Protein Acetylation in Diverse Biological Processes in Rice (Oryza sativa)

    PubMed Central

    Zhong, Xiaoxian; Tan, Feng; Mujahid, Hana; Zhang, Jian; Nanduri, Bindu; Peng, Zhaohua

    2014-01-01

    Lysine acetylation is a reversible, dynamic protein modification regulated by lysine acetyltransferases and deacetylases. Recent advances in high-throughput proteomics have greatly contributed to the success of global analysis of lysine acetylation. A large number of proteins of diverse biological functions have been shown to be acetylated in several reports in human cells, E.coli, and dicot plants. However, the extent of lysine acetylation in non-histone proteins remains largely unknown in monocots, particularly in the cereal crops. Here we report the mass spectrometric examination of lysine acetylation in rice (Oryza sativa). We identified 60 lysine acetylated sites on 44 proteins of diverse biological functions. Immunoblot studies further validated the presence of a large number of acetylated non-histone proteins. Examination of the amino acid composition revealed substantial amino acid bias around the acetylation sites and the amino acid preference is conserved among different organisms. Gene ontology analysis demonstrates that lysine acetylation occurs in diverse cytoplasmic, chloroplast and mitochondrial proteins in addition to the histone modifications. Our results suggest that lysine acetylation might constitute a regulatory mechanism for many proteins, including both histones and non-histone proteins of diverse biological functions. PMID:24586658

  18. Methionine as a potential precursor for halogenated compounds by the reaction with iron minerals

    NASA Astrophysics Data System (ADS)

    Tubbesing, C.; Krause, T.; Mulder, I.; Kotte, K.; Schöler, H. F.

    2012-04-01

    Volatile halogenated compounds (VOX) play an important role in different photochemical reactions within the troposphere and the stratosphere. Soils and sediments seem to act as a major natural source for VOX, but investigations of the reaction mechanisms are rather scarce. To get further information on potential intermediates the reaction of the amino acid methionine with the ferrous and ferric iron minerals pyrite and ferrihydrite as well as solute ferrous sulfate was studied using a gas chromatography-flame ionization detector (GC-FID). Methionine is an important amino acid in the biosynthesis of plants used as a starting compound for the messenger ethene with aminocyclopropane carboxylic acid as an intermediate product. This pathway may also occur under abiotic conditions. Ethene is assumed as precursor for various halogenated C2-compounds like vinyl chloride and dichloroethene. Due to its ubiquity by an average concentration of 10 to 290 ng/g soil and its potential to regenerate in soils and organic litter by microorganisms, methionine may be an important educt for both abiotic and biotic terrestrial halogenation processes. In laboratory tests methionine was exposed to different iron species like pyrite, iron sulfate or ferrihydrite. The oxidant H2O2 was used to start the reaction. Production values of methyl chloride and other halogenated compounds are discussed in the context of methionine as their potential precursor and several Fe-minerals as soil-borne catalysers. Several possible intermediates for the production of VOX have been detected e.g. methane, ethene or propane. A formation of isobutylene is noteworthy for some cases. In addition to VOC the production of methyl chloride and dimethyl sulfide (DMS) was observed. Only the DMS bears upon a specific mineral. The samples containing pyrite reveal the highest concentrations. To get a better assessment of methionine, respectively VOC released from methionine as precursors for halogenated compounds

  19. Dysregulated Hepatic Methionine Metabolism Drives Homocysteine Elevation in Diet-Induced Nonalcoholic Fatty Liver Disease.

    PubMed

    Pacana, Tommy; Cazanave, Sophie; Verdianelli, Aurora; Patel, Vaishali; Min, Hae-Ki; Mirshahi, Faridoddin; Quinlivan, Eoin; Sanyal, Arun J

    2015-01-01

    methyltransferase Dnmt3a decreased, the global DNA methylation was unaltered. Among individual genes, only HMG-CoA reductase (Hmgcr) was hypermethylated, and no methylation changes were observed in fatty acid synthase (Fasn), nuclear factor of kappa light polypeptide gene enhancer in B-cells 1 (Nfκb1), c-Jun, B-cell lymphoma 2 (Bcl-2) and Caspase 3. NAFLD was associated with hepatic methionine deficiency and homocysteine elevation, resulting mainly from impaired homocysteine remethylation, and aberrancy in methyltransferase reactions. Despite increased PRMT1 expression, hepatic ADMA was depleted while circulating ADMA was increased, suggesting increased export to circulation.

  20. Hemoglobin Labeled by Radioactive Lysine

    DOE R&D Accomplishments Database

    Bale, W. F.; Yuile, C. L.; DeLaVergne, L.; Miller, L. L.; Whipple, G. H.

    1949-12-08

    This paper reports on the utilization of tagged epsilon carbon of DL-lysine by a dog both anemic and hypoproteinemic due to repeated bleeding plus a diet low in protein. The experiment extended over period of 234 days, a time sufficient to indicate an erythrocyte life span of at least 115 days based upon the rate of replacement of labeled red cell proteins. The proteins of broken down red cells seem not to be used with any great preference for the synthesis of new hemoglobin.

  1. Reconstruction of Cysteine Biosynthesis Using Engineered Cysteine-Free and Methionine-Free Enzymes

    NASA Technical Reports Server (NTRS)

    Wang, Kendrick; Fujishima, Kosuke; Abe, Nozomi; Nakahigashi, Kenji; Endy, Drew; Rothschild, Lynn J.

    2016-01-01

    Ten of the proteinogenic amino acids can be generated abiotically while the remaining thirteen require biology for their synthesis. Paradoxically, the biosynthesis pathways observed in nature require enzymes that are made with the amino acids they produce. For example, Escherichia coli produces cysteine from serine via two enzymes that contain cysteine. Here, we substituted alternate amino acids for cysteine and also methionine, which is biosynthesized from cysteine, in serine acetyl transferase (CysE) and O-acetylserine sulfhydrylase (CysM). CysE function was rescued by cysteine-and-methionine-free enzymes and CysM function was rescued by cysteine-free enzymes. Structural modeling suggests that methionine stabilizes CysM and is present in the active site of CysM. Cysteine is not conserved among CysE and CysM protein orthologs, suggesting that cysteine is not functionally important for its own synthesis. Engineering biosynthetic enzymes that lack the amino acids being synthesized provides insights into the evolution of amino acid biosynthesis and pathways for bioengineering.

  2. Genome-Wide Analysis of the Lysine Biosynthesis Pathway Network during Maize Seed Development

    PubMed Central

    Liu, Yuwei; Xie, Shaojun; Yu, Jingjuan

    2016-01-01

    Lysine is one of the most limiting essential amino acids for humans and livestock. The nutritional value of maize (Zea mays L.) is reduced by its poor lysine content. To better understand the lysine biosynthesis pathway in maize seed, we conducted a genome-wide analysis of the genes involved in lysine biosynthesis. We identified lysine biosynthesis pathway genes (LBPGs) and investigated whether a diaminopimelate pathway variant exists in maize. We analyzed two genes encoding the key enzyme dihydrodipicolinate synthase, and determined that they contribute differently to lysine synthesis during maize seed development. A coexpression network of LBPGs was constructed using RNA-sequencing data from 21 developmental stages of B73 maize seed. We found a large set of genes encoding ribosomal proteins, elongation factors and zein proteins that were coexpressed with LBPGs. The coexpressed genes were enriched in cellular metabolism terms and protein related terms. A phylogenetic analysis of the LBPGs from different plant species revealed different relationships. Additionally, six transcription factor (TF) families containing 13 TFs were identified as the Hub TFs of the LBPGs modules. Several expression quantitative trait loci of LBPGs were also identified. Our results should help to elucidate the lysine biosynthesis pathway network in maize seed. PMID:26829553

  3. Synthesis and Characterization of L-Lysine Conjugated Silver Nanoparticles Smaller Than 10 nM

    PubMed Central

    Bonor, Jeremy; Reddy, Vandhana; Akkiraju, Hemanth; Dhurjati, Prasad; Nohe, Anja

    2015-01-01

    A rapid and convenient batch method for synthesizing lysine-conjugated silver nanoparticles of approximately 5 nm of size was developed. Nanoparticles of size less than 100 nm exhibit significant medical potential. L-Lysine demonstrates potential for therapeutic applications and silver nanoparticles are an optimal choice for drug delivery because of its intrinsic anti-platelet, anti-bacterial and anti-inflammatory capabilities. Current synthesis protocols for Lysine-capped particles under 10 nm are time consuming and tedious and allow only for the sythesis of small quantities of particles. The synthesis of Lysin-capped silver nanoparticles was based on the reaction in which AgNO3 was reduced by excess NaBH4. L-Lysine, a known essential amino acid, served as the capping agent to minimize initial aggregation. The particles were then separated by size chromatography. Capping occurred through the amide bond on L-Lysine as determined by FT-IR. The conjugation of the particle to the amide bond is important, since this leaves the amino group of Lysine open to further modifications. The particles were further characterized in regards to their shape, size and stability. Finally we demonstrated that the synthesized particles exhibit limited to no toxicity in cells, using HEK 293 cell line as a model system. Our sythesis protocol can be successfully used for scale-up and synthesis of high quantities of nanoparticles. PMID:26478827

  4. Structural Basis for l-Lysine Feedback Inhibition of Homocitrate Synthase

    SciTech Connect

    Bulfer, Stacie L.; Scott, Erin M.; Pillus, Lorraine; Trievel, Raymond C.

    2010-09-02

    The {alpha}-aminoadipate pathway of lysine biosynthesis is modulated at the transcriptional and biochemical levels by feedback inhibition. The first enzyme in the {alpha}-aminoadipate pathway, homocitrate synthase (HCS), is the target of the feedback regulation and is strongly inhibited by L-lysine. Here we report the structure of Schizosaccharomyces pombe HCS (SpHCS) in complex with L-lysine. The structure illustrates that the amino acid directly competes with the substrate 2-oxoglutarate for binding within the active site of HCS. Differential recognition of the substrate and inhibitor is achieved via a switch position within the ({alpha}/{beta}){sub 8} TIM barrel of the enzyme that can distinguish between the C5-carboxylate group of 2-oxoglutarate and the {epsilon}-ammonium group of L-lysine. In vitro and in vivo assays demonstrate that mutations of the switch residues, which interact with the L-lysine {epsilon}-ammonium group, abrogate feedback inhibition, as do substitutions of residues within the C-terminal domain that were identified in a previous study of L-lysine-insensitive HCS mutants in Saccharomyces cerevisiae. Together, these results yield new insights into the mechanism of feedback regulation of an enzyme central to lysine biosynthesis.

  5. Effect of different postharvest temperatures on the accumulation of sugars, organic acids, and amino acids in the juice sacs of Satsuma mandarin (Citrus unshiu Marc.) fruit.

    PubMed

    Matsumoto, Hikaru; Ikoma, Yoshinori

    2012-10-03

    To elucidate the effect of different postharvest temperatures on the accumulation of sugars, organic acids, and amino acids and to determine the best temperature to minimize their postharvest change, their content after harvest was investigated at 5, 10, 20, and 30 °C for 14 days in the juice sacs of Satsuma mandarin (Citrus unshiu Marc. cv. Aoshima-unshiu) fruit. In all sugars, the changes were negligible at all temperatures. Organic acids decreased slightly at all temperatures, with the exception of malic acid at 30 °C, which increased slightly. Two amino acids, ornithine and glutamine, increased at 5 °C, but they did not increase at other temperatures. In 11 amino acids (phenylalanine, tryptophan, tyrosine, isoleucine, leucine, valine, threonine, lysine, methionine, histidine, and γ-amino butyric acid), the content was higher at 20 and 30 °C than at other temperatures. Thus, the content of amino acids was more variable than that of sugars and organic acids in response to temperatures. Moreover, amino acids responded to temperature differently: two amino acids were cold responsive, and 11 were heat-responsive. The best temperature to minimize the postharvest changes in amino acid profiles in the juice sacs of Aoshima-unshiu was 10 °C. The responsiveness to temperatures in two cold-responsive (ornithine and glutamine) and five heat-responsive (phenylalanine, tryptophan, valine, lysine, and histidine) amino acids was conserved among three different Satsuma mandarin cultivars, Aoshima-unshiu (late-maturing cultivar), Silverhill (midmaturing cultivar), and Miyagawa-wase (early-maturing cultivar). The metabolic responsiveness to temperature stress was discussed on the basis of the changes in the amino acid profile.

  6. Methionine restriction beyond life-span extension.

    PubMed

    Ables, Gene P; Hens, Julie R; Nichenametla, Sailendra N

    2016-01-01

    Dietary methionine restriction (MR) extends life span across species via various intracellular regulatory mechanisms. In rodents, MR induces resistance against adiposity, improves hepatic glucose metabolism, preserves cardiac function, and reduces body size, all of which can affect the onset of age-related diseases. Recent studies have shown that MR-affected biomarkers, such as fibroblast growth factor 21, adiponectin, leptin, cystathionine β synthase, and insulin-like growth factor 1, can potentially alter physiology. The beneficial effects of MR could be explained in part by its ability to reduce mitochondrial oxidative stress. Studies have revealed that MR can reduce reactive oxygen species that damage cells and promote cancer progression. It has been demonstrated that either MR or the targeting of specific genes in the methionine cycle could induce cell apoptosis while decreasing proliferation in several cancer models. The complete mechanism underlying the actions of MR on the cell cycle during cancer has not been fully elucidated. Epigenetic mechanisms, such as methylation and noncoding RNAs, are also possible downstream effectors of MR; future studies should help to elucidate some of these mechanisms. Despite evidence that changes in dietary methionine can affect epigenetics, it remains unknown whether epigenetics is a mechanism in MR. This review summarizes research on MR and its involvement in metabolism, cancer, and epigenetics.

  7. Transcriptional upregulation of four genes of the lysine biosynthetic pathway by homocitrate accumulation in Penicillium chrysogenum: homocitrate as a sensor of lysine-pathway distress.

    PubMed

    Teves, Franco; Lamas-Maceiras, Mónica; García-Estrada, Carlos; Casqueiro, Javier; Naranjo, Leopoldo; Ullán, Ricardo V; Scervino, José-Martín; Wu, Xiaobin; Velasco-Conde, Tania; Martín, Juan F

    2009-12-01

    The lysine biosynthetic pathway has to supply large amounts of alpha-aminoadipic acid for penicillin biosynthesis in Penicillium chrysogenum. In this study, we have characterized the P. chrysogenum L2 mutant, a lysine auxotroph that shows highly increased expression of several lysine biosynthesis genes (lys1, lys2, lys3, lys7). The L2 mutant was found to be deficient in homoaconitase activity since it was complemented by the Aspergillus nidulans lysF gene. We have cloned a gene (named lys3) that complements the L2 mutation by transformation with a P. chrysogenum genomic library, constructed in an autonomous replicating plasmid. The lys3-encoded protein showed high identity to homoaconitases. In addition, we cloned the mutant lys3 allele from the L2 strain that showed a G(1534) to A(1534) point mutation resulting in a Gly(495) to Asp(495) substitution. This mutation is located in a highly conserved region adjacent to two of the three cysteine residues that act as ligands to bind the iron-sulfur cluster required for homoaconitase activity. The L2 mutant accumulates homocitrate. Deletion of the lys1 gene (homocitrate synthase) in the L2 strain prevented homocitrate accumulation and reverted expression levels of the four lysine biosynthesis genes tested to those of the parental prototrophic strain. Homocitrate accumulation seems to act as a sensor of lysine-pathway distress, triggering overexpression of four of the lysine biosynthesis genes.

  8. Reinventing cell penetrating peptides using glycosylated methionine sulfonium ion sequences

    DOE PAGES

    Kramer, Jessica R.; Schmidt, Nathan W.; Mayle, Kristine M.; ...

    2015-04-15

    Cell penetrating peptides (CPPs) are intriguing molecules that have received much attention, both in terms of mechanistic analysis and as transporters for intracellular therapeutic delivery. Most CPPs contain an abundance of cationic charged residues, typically arginine, where the amino acid compositions, rather than specific sequences, tend to determine their ability to enter cells. Hydrophobic residues are often added to cationic sequences to create efficient CPPs, but typically at the penalty of increased cytotoxicity. Here, we examined polypeptides containing glycosylated, cationic derivatives of methionine, where we found these hydrophilic polypeptides to be surprisingly effective as CPPs and to also possess lowmore » cytotoxicity. X-ray analysis of how these new polypeptides interact with lipid membranes revealed that the incorporation of sterically demanding hydrophilic cationic groups in polypeptides is an unprecedented new concept for design of potent CPPs.« less

  9. Reinventing cell penetrating peptides using glycosylated methionine sulfonium ion sequences

    SciTech Connect

    Kramer, Jessica R.; Schmidt, Nathan W.; Mayle, Kristine M.; Kamei, Daniel T.; Wong, Gerard C.L.; Deming, Timothy J.

    2015-04-15

    Cell penetrating peptides (CPPs) are intriguing molecules that have received much attention, both in terms of mechanistic analysis and as transporters for intracellular therapeutic delivery. Most CPPs contain an abundance of cationic charged residues, typically arginine, where the amino acid compositions, rather than specific sequences, tend to determine their ability to enter cells. Hydrophobic residues are often added to cationic sequences to create efficient CPPs, but typically at the penalty of increased cytotoxicity. Here, we examined polypeptides containing glycosylated, cationic derivatives of methionine, where we found these hydrophilic polypeptides to be surprisingly effective as CPPs and to also possess low cytotoxicity. X-ray analysis of how these new polypeptides interact with lipid membranes revealed that the incorporation of sterically demanding hydrophilic cationic groups in polypeptides is an unprecedented new concept for design of potent CPPs.

  10. Reinventing Cell Penetrating Peptides Using Glycosylated Methionine Sulfonium Ion Sequences

    PubMed Central

    2015-01-01

    Cell penetrating peptides (CPPs) are intriguing molecules that have received much attention, both in terms of mechanistic analysis and as transporters for intracellular therapeutic delivery. Most CPPs contain an abundance of cationic charged residues, typically arginine, where the amino acid compositions, rather than specific sequences, tend to determine their ability to enter cells. Hydrophobic residues are often added to cationic sequences to create efficient CPPs, but typically at the penalty of increased cytotoxicity. Here, we examined polypeptides containing glycosylated, cationic derivatives of methionine, where we found these hydrophilic polypeptides to be surprisingly effective as CPPs and to also possess low cytotoxicity. X-ray analysis of how these new polypeptides interact with lipid membranes revealed that the incorporation of sterically demanding hydrophilic cationic groups in polypeptides is an unprecedented new concept for design of potent CPPs. PMID:27162954

  11. The lysP gene encodes the lysine-specific permease.

    PubMed Central

    Steffes, C; Ellis, J; Wu, J; Rosen, B P

    1992-01-01

    Escherichia coli transports lysine by two distinct systems, one of which is specific for lysine (LysP) and the other of which is inhibited by arginine ornithine. The activity of the lysine-specific system increases with growth in acidic medium, anaerobiosis, and high concentrations of lysine. It is inhibited by the lysine analog S-(beta-aminoethyl)-L-cysteine (thiosine). Thiosine-resistant (Tsr) mutants were isolated by using transpositional mutagenesis with TnphoA. A Tsr mutant expressing alkaline phosphatase activity in intact cells was found to lack lysine-specific transport. This lysP mutation was mapped to about 46.5 min on the E. coli chromosome. The lysP-phoA fusion was cloned and used as a probe to clone the wild-type lysP gene. The nucleotide sequence of the 2.7-kb BamHI fragment was determined. An open reading frame from nucleotides 522 to 1989 was observed. The translation product of this open reading frame is predicted to be a hydrophobic protein of 489 residues. The lysP gene product exhibits sequence similarity to a family of amino acid transport proteins found in both prokaryotes and eukaryotes, including the aromatic amino acid permease of E. coli (aroP) and the arginine permease of Saccharomyces cerevisiae (CAN1). Cells carrying a plasmid with the lysP gene exhibited a 10- to 20-fold increase in the rate of lysine uptake above wild-type levels. These results demonstrate that the lysP gene encodes the lysine-specific permease. Images PMID:1315732

  12. Dietary L-lysine prevents arterial calcification in adenine-induced uremic rats.

    PubMed

    Shimomura, Akihiro; Matsui, Isao; Hamano, Takayuki; Ishimoto, Takuya; Katou, Yumiko; Takehana, Kenji; Inoue, Kazunori; Kusunoki, Yasuo; Mori, Daisuke; Nakano, Chikako; Obi, Yoshitsugu; Fujii, Naohiko; Takabatake, Yoshitsugu; Nakano, Takayoshi; Tsubakihara, Yoshiharu; Isaka, Yoshitaka; Rakugi, Hiromi

    2014-09-01

    Vascular calcification (VC) is a life-threatening complication of CKD. Severe protein restriction causes a shortage of essential amino acids, and exacerbates VC in rats. Therefore, we investigated the effects of dietary l-lysine, the first-limiting amino acid of cereal grains, on VC. Male Sprague-Dawley rats at age 13 weeks were divided randomly into four groups: low-protein (LP) diet (group LP), LP diet+adenine (group Ade), LP diet+adenine+glycine (group Gly) as a control amino acid group, and LP diet+adenine+l-lysine·HCl (group Lys). At age 18 weeks, group LP had no VC, whereas groups Ade and Gly had comparable levels of severe VC. l-Lysine supplementation almost completely ameliorated VC. Physical parameters and serum creatinine, urea nitrogen, and phosphate did not differ among groups Ade, Gly, and Lys. Notably, serum calcium in group Lys was slightly but significantly higher than in groups Ade and Gly. Dietary l-lysine strongly suppressed plasma intact parathyroid hormone in adenine rats and supported a proper bone-vascular axis. The conserved orientation of the femoral apatite in group Lys also evidenced the bone-protective effects of l-lysine. Dietary l-lysine elevated plasma alanine, proline, arginine, and homoarginine but not lysine. Analyses in vitro demonstrated that alanine and proline inhibit apoptosis of cultured vascular smooth muscle cells, and that arginine and homoarginine attenuate mineral precipitations in a supersaturated calcium/phosphate solution. In conclusion, dietary supplementation of l-lysine ameliorated VC by modifying key pathways that exacerbate VC.

  13. Dietary l-Lysine Prevents Arterial Calcification in Adenine-Induced Uremic Rats

    PubMed Central

    Shimomura, Akihiro; Matsui, Isao; Hamano, Takayuki; Ishimoto, Takuya; Katou, Yumiko; Takehana, Kenji; Inoue, Kazunori; Kusunoki, Yasuo; Mori, Daisuke; Nakano, Chikako; Obi, Yoshitsugu; Fujii, Naohiko; Takabatake, Yoshitsugu; Nakano, Takayoshi; Tsubakihara, Yoshiharu; Rakugi, Hiromi

    2014-01-01

    Vascular calcification (VC) is a life-threatening complication of CKD. Severe protein restriction causes a shortage of essential amino acids, and exacerbates VC in rats. Therefore, we investigated the effects of dietary l-lysine, the first-limiting amino acid of cereal grains, on VC. Male Sprague-Dawley rats at age 13 weeks were divided randomly into four groups: low-protein (LP) diet (group LP), LP diet+adenine (group Ade), LP diet+adenine+glycine (group Gly) as a control amino acid group, and LP diet+adenine+l-lysine·HCl (group Lys). At age 18 weeks, group LP had no VC, whereas groups Ade and Gly had comparable levels of severe VC. l-Lysine supplementation almost completely ameliorated VC. Physical parameters and serum creatinine, urea nitrogen, and phosphate did not differ among groups Ade, Gly, and Lys. Notably, serum calcium in group Lys was slightly but significantly higher than in groups Ade and Gly. Dietary l-lysine strongly suppressed plasma intact parathyroid hormone in adenine rats and supported a proper bone-vascular axis. The conserved orientation of the femoral apatite in group Lys also evidenced the bone-protective effects of l-lysine. Dietary l-lysine elevated plasma alanine, proline, arginine, and homoarginine but not lysine. Analyses in vitro demonstrated that alanine and proline inhibit apoptosis of cultured vascular smooth muscle cells, and that arginine and homoarginine attenuate mineral precipitations in a supersaturated calcium/phosphate solution. In conclusion, dietary supplementation of l-lysine ameliorated VC by modifying key pathways that exacerbate VC. PMID:24652795

  14. Volatile sulphur compounds and pathways of L-methionine catabolism in Williopsis yeasts.

    PubMed

    Tan, Amelia W J; Lee, Pin-Rou; Seow, Yi-Xin; Ong, Peter K C; Liu, Shao-Quan

    2012-08-01

    Volatile sulphur compounds (VSCs) are important to the food industry due to their high potency and presence in many foods. This study assessed for the first time VSC production and pathways of L: -methionine catabolism in yeasts from the genus Williopsis with a view to understanding VSC formation and their potential flavour impact. Five strains of Williopsis saturnus (var. saturnus, var. subsufficiens, var. suavolens, var. sargentensis and var. mrakii) were screened for VSC production in a synthetic medium supplemented with L: -methionine. A diverse range of VSCs were produced including dimethyl disulphide, dimethyl trisulphide, 3-(methylthio)-1-propanal (methional), 3-(methylthio)-1-propanol (methionol), 3-(methylthio)-1-propene, 3-(methylthio)-1-propyl acetate, 3-(methylthio)-1-propanoic acid (methionic acid) and ethyl 3-(methylthio)-1-propanoate, though the production of these VSCs varied between yeast strains. W. saturnus var. saturnus NCYC22 was selected for further studies due to its relatively high VSC production. VSC production was characterised step-wise with yeast strain NCYC22 in coconut cream at different L: -methionine concentrations (0.00-0.20%) and under various inorganic sulphate (0.00-0.20%) and nitrogen (ammonia) supplementation (0.00-0.20%), respectively. Optimal VSC production was obtained with 0.1% of L: -methionine, while supplementation of sulphate had no significant effect. Nitrogen supplementation showed a dramatic inhibitory effect on VSC production. Based on the production of VSCs, the study suggests that the Ehrlich pathway of L: -methionine catabolism is operative in W. saturnus yeasts and can be manipulated by adjusting certain nutrient parameters to control VSC production.

  15. Standardized ileal digestibility of proteins and amino acids in sesame expeller and soya bean meal in weaning piglets.

    PubMed

    Aguilera, A; Reis de Souza, T C; Mariscal-Landín, G; Escobar, K; Montaño, S; Bernal, M G

    2015-08-01

    Apparent ileal digestibility (AID) of diets containing sesame expeller (SE) and soya bean meal (SBM) was determined using 15 piglets (Genetiporc(®)), weaned at 17 ± 0.4 days with average body weight of 6.4 ± 0.7 kg (Fertilis 20 × G Performance, Genetiporc(®), PIC México, Querétaro, México). Piglets were randomly assigned to three treatments: (i) a reference diet with casein as the sole protein source; (ii) a mixed diet of casein-SE; and (iii) a mixed diet of casein-SBM. The chemical composition of SE and SBM was determined, and AID and standardized ileal digestibility (SID) of crude protein (CP) and amino acids (AAs) were determined for each protein source. SE contained greater quantities of ether extract, neutral detergent fibre, phytic acid, methionine and arginine than SBM. Lysine and proline contents and trypsin inhibitor activity were higher in SBM than in SE. The AID and SID of CP and AA (except for lysine and proline) were similar in SE and SBM. The AID of lysine and proline was higher in SBM than in SE (p < 0.05), and the SID of proline was higher in SE than in SBM (p < 0.05). These findings indicate that SE is an appropriate alternative protein source for early weaned pigs.

  16. Comparison of the effects of seleno-l-methionine, seleno-dl-methionine, and selenized yeast on reproduction of mallards

    USGS Publications Warehouse

    Heinz, G.H.; Hoffman, D.J.

    1996-01-01

    The toxicities of seleno-L-methionine, seleno-DL-methionine, and selenized yeast were compared. Ten pairs of mallards were fed a control diet and 15 pairs were fed diets containing 10 ppm selenium as seleno-DL-methionine, seleno-L-methionine, or selenized yeast. Hatching of fertile eggs was significantly lower for females fed 10 ppm selenium as seleno-DL-methionine (7.6%) and seleno-L-methionine (6.4%) than for controls (41.3%). Survival of ducklings was lower when their parents had been fed 10 ppm selenium as seleno-L-methionine (20.0%) than for controls (98.4%). The number of 6-day-old ducklings produced per female was significantly lower for mallards fed 10 ppm selenium as seleno-DL-methionine (0.47) or selenized yeast (2.67) than for controls (6.10), and was significantly lower for mallards fed seleno-L-methionine (0.13) than for mallards fed selenized yeast. The eighth eggs of females fed the DL or L forms of selenomethionine contained means of 9.2 and 8.9 ppm selenium, wet weight; these means were higher than the mean (6.6 ppm) for females fed selenized yeast. Among embryos that died at 7 days of age or older, the percentage of embryos that were deformed was 1.3% for controls, 24.6% for seleno-DL-methionine, 28.2% for seleno-L-methionine, and 11.0% for selenized yeast. The results suggested that seleno-DL-methionine and seleno-L-methionine were of similar toxicity and were both more toxic than selenium from selenized yeast.

  17. Carbon-11-methionine and PET in evaluation of treatment response of breast cancer.

    PubMed Central

    Huovinen, R.; Leskinen-Kallio, S.; Någren, K.; Lehikoinen, P.; Ruotsalainen, U.; Teräs, M.

    1993-01-01

    Uptake of L-methyl-11C-methionine (11C-methionine) in breast cancer metastases was studied with positron emission tomography (PET). Eight patients with soft tissue metastases were studied twice: before the onset of chemotherapy (4), hormonal therapy (3) or radiotherapy (1) and 3-14 weeks later. The radioactivity concentration of the low molecular weight fraction of venous plasma samples separated by fast gel filtration was used as input function. The input corrected uptake rate of 11C-methionine (Ki) in breast cancer metastases before the treatment ranged between 0.035 and 0.186 1 min-1 and the standardised uptake value (SUV) between 2.0 and 11.4. The uptake of 11C-methionine into the metastases decreased when clinical objective stability or regression of the metastases was later obtained and increased in cases where progressive disease was seen during treatment. We conclude that metabolic changes in the amino acid metabolism detected by PET precede the clinical response, and may be of clinical value in predicting the treatment response. Images Figure 1 PMID:8471437

  18. Characteristics of viable Brevibacterium linens cells, methionine and cysteine in milkfat-coated microcapsules.

    PubMed

    Kim, S C; Olson, N F

    1985-01-01

    The potential for microencapsulation of viable Brevibacterium linens with methionine or cysteine in milkfat to produce sulphur compounds was examined in this study. More than 80 per cent of B. linens cells were encapsulated and then numbers inside capsules increased about three-fold during 48 h at 26 degrees C under anaerobic conditions before a slow decline. Most of micro-organisms (7 x 10(7)/ml) were still viable in the capsules after 15 days. More than 90 per cent of cysteine and methionine were encapsulated and 90, 80 and 60 per cent of the encapsulated amino acids were maintained in the capsules at 4, 12 and 26 degrees C, respectively, after 24 h. Most of the cysteine was oxidized to cystine during microencapsulation but still available to micro-organisms whereas methionine remained in the reduced form. Partition coefficients of methionine and cysteine to milkfat were 0.117 and 0.275, respectively, indicating that most of these substrates would be available to cells of B. linens in the capsules.

  19. Lysine Transport across Isolated Rabbit Ileum

    PubMed Central

    Munck, B. G.; Schultz, Stanley G.

    1969-01-01

    Lysine transport by in vitro distal rabbit ileum has been investigated by determining (a) transmural fluxes across short-circuited segments of the tissue; (b) accumulation by mucosal strips; and (c) influx from the mucosal solution across the brush border into the epithelium. Net transmural flux of lysine is considerably smaller than that of alanine. However, lysine influx across the brush border and lysine accumulation by mucosal strips are quantitatively comparable to alanine influx and accumulation. Evidence is presented that the "low transport capacity" of rabbit ileum for lysine is due to: (a) a carrier-mediated process responsible for efflux of lysine out of the cell across the serosal and/or lateral membranes that is characterized by a low maximal velocity; and (b) a high "backflux" of lysine out of the cell across the mucosal membrane. A possible explanation for the latter observation is discussed with reference to the relatively low Na dependence of lysine transport across the intestinal brush border. PMID:5764744

  20. Stimulation of Ethylene Production in Apple Tissue Slices by Methionine

    PubMed Central

    Lieberman, Morris; Kunishi, Alice; Mapson, L. W.; Wardale, D. A.

    1966-01-01

    Methionine can induce more than a 100% increase in ethylene production by apple tissue slices. The increased amount of ethylene derives from carbons 3 and 4 of methionine. Only post-climacteric fruit tissues are stimulated by methionine, and stimulation is optimum after 8 months' storage. Copper chelators such as sodium diethyl dithiocarbamate and cuprizone very markedly inhibit ethylene production by tissue slices. Carbon monoxide does not effect ethylene production by the slices. These data suggest that the mechanism for the conversion of methionine to ethylene, in apple tissues, is similar to the previously described model system for producing ethylene from methionine and reduced copper. Therefore, it is suggested that one of the ethylene-forming systems in tissues derives from methionine and proceeds to ethylene via a copper enzyme system which may be a peroxidase. PMID:16656267

  1. Comparative dynamics of methionine side-chain in FMOC-methionine and in amyloid fibrils

    NASA Astrophysics Data System (ADS)

    Vugmeyster, Liliya; Ostrovsky, Dmitry

    2017-04-01

    We compared the dynamics of key methionine methyl groups in the water-accessible hydrophobic cavity of amyloid fibrils and Fluorenylmethyloxycarbonyl-Methionine (FMOC-Met), which renders general hydrophobicity to the environment without the complexity of the protein. Met35 in the hydrated cavity was recently found to undergo a dynamical cross-over from the dominance of methyl rotations at low temperatures to the dominance of diffusive motion of methyl axis at high temperatures. Current results indicate that in FMOC-Met this cross-over is suppressed, similar to what was observed for the dry fibrils, indicating that hydration of the cavity is driving the onset of the dynamical transition.

  2. The Construction and Expression of Lysine-Rich Gene in the Mammary Gland of Transgenic Mice

    PubMed Central

    Ma, Xin; Zhang, Peng; Song, Guangqi; Chen, Yue; Wang, Zhongwei; Yin, Yupeng; Kong, Delong; Zhang, Sheng; Zhao, Zhihui; Ouyang, Hongsheng

    2012-01-01

    Lysine is the limiting amino acid in cereal grains, which represent a major source of human food and animal feed worldwide, and is considered the most important of the essential amino acids. In this study, β-casein, αS2-casein, and lactotransferrin cDNA clone fragments encoding lysine-rich peptides were fused together to generate a lysine-rich (LR) gene and the mammary gland-specific expression vector pBC1-LR-NEOr was constructed. Transgenic mice were generated by pronuclear microinjection of the linearized expression vectors harboring the LR transgene. The transgenic mice and their offspring were examined using multiplex polymerase chain reaction (PCR), Southern blotting, reverse transcriptase–PCR, in situ hybridization, and Western blotting techniques. Our results showed that the LR gene was successfully integrated into the mouse genome and was transmitted stably. The specific LR gene expression was restricted to the mammary gland, active alveoli of the transgenic female mice during lactation. The lysine level of the two transgenic lines was significantly higher than that of nontransgenic controls (p<0.05). In addition, the growth performance of transgenic pups was enhanced by directly feeding them the LR protein-enriched transgenic milk. Our results demonstrated that lysine-rich gene was successfully constructed and expressed in mammary gland of transgenic mice. This study will provide a better understanding of how mammary gland expression systems that increase the lysine content of milk can be applied to other mammals, such as cows. PMID:22577831

  3. Na/sup +/-dependent transport of /sup 14/C-L-lysine across bullfrog alveolar epithelium

    SciTech Connect

    Kim, K.J.; Crandall, E.D.

    1986-03-01

    Transepithelial transport of the basic amino acid L-lysine has been studied utilizing the isolated intact bullfrog lung mounted in the Ussing chamber. Lungs were excised from doubly pithed bullfrogs and sandwiched between two hemichambers. /sup 14/C-(U)-L-lysine was added to the upstream reservoir of amphibian Ringer solution, while the tissue was short-circuited. Two lungs from the same animal were used simultaneously to determine the two opposite unidirectional fluxes. Downstream and upstream radioactivities were assayed and used to estimate the apparent permeability (P) of the labeled lysine. Results indicate that the apparent P of /sup 14/C-L-lysine measured in the alveolar (M) to the pleural (S) direction is 19.06 (+- 2.84) x 10/sup -7/ cm/s and P in the S to M direction is 3.29 (+- 0.02) x 10/sup -7/ cm/s. When the 100 mM NaCl in the bath was replaced by 110 mM choline chloride, the flux of /sup 14/C-L-lysine from the alveolar to the pleural side decreased to the same value as that in the opposite direction. The flux from the pleural to the alveolar direction in the absence of Na/sup +/ did not change. These results suggest that the alveolar epithelium exhibits Na/sup +/-dependent amino acid (L-lysine) transport in the M->S, but not in the S->M, direction.

  4. The construction and expression of lysine-rich gene in the mammary gland of transgenic mice.

    PubMed

    Ma, Xin; Zhang, Peng; Song, Guangqi; Chen, Yue; Wang, Zhongwei; Yin, Yupeng; Kong, Delong; Zhang, Sheng; Zhao, Zhihui; Ouyang, Hongsheng; Tang, Bo; Li, Ziyi

    2012-08-01

    Lysine is the limiting amino acid in cereal grains, which represent a major source of human food and animal feed worldwide, and is considered the most important of the essential amino acids. In this study, β-casein, αS2-casein, and lactotransferrin cDNA clone fragments encoding lysine-rich peptides were fused together to generate a lysine-rich (LR) gene and the mammary gland-specific expression vector pBC1-LR-NEO(r) was constructed. Transgenic mice were generated by pronuclear microinjection of the linearized expression vectors harboring the LR transgene. The transgenic mice and their offspring were examined using multiplex polymerase chain reaction (PCR), Southern blotting, reverse transcriptase-PCR, in situ hybridization, and Western blotting techniques. Our results showed that the LR gene was successfully integrated into the mouse genome and was transmitted stably. The specific LR gene expression was restricted to the mammary gland, active alveoli of the transgenic female mice during lactation. The lysine level of the two transgenic lines was significantly higher than that of nontransgenic controls (p<0.05). In addition, the growth performance of transgenic pups was enhanced by directly feeding them the LR protein-enriched transgenic milk. Our results demonstrated that lysine-rich gene was successfully constructed and expressed in mammary gland of transgenic mice. This study will provide a better understanding of how mammary gland expression systems that increase the lysine content of milk can be applied to other mammals, such as cows.

  5. One-pot modification of 5'-capped RNA based on methionine analogs.

    PubMed

    Muttach, Fabian; Rentmeister, Andrea

    2016-09-01

    This paper outlines chemically and enzymatically synthesized S-adenosylmethionine (AdoMet) analogs and their use in the site-specific modification of RNA by methyltransferases, enabling the facile attachment of clickable moieties to the nucleic acid. We then focus on methodological aspects of setting up a methyltransferase-based enzymatic cascade reaction starting from methionine analogs. This strategy is applied to the one-pot modification of the mRNA cap which is subsequently derivatized in copper-free and copper-catalyzed click reactions. We show that high transfer efficiencies to the cap are obtained using Se-propargyl-, hexenynyl- and azido-bearing methionine analogs. By switching to other methyltransferases our one-pot modification approach should be directly applicable to the regiospecific modification of other target molecules including nucleic acids, proteins and small molecules.

  6. Isolation and primary structure of a methionine- and cystine-rich seed protein of Cannabis sativa.

    PubMed

    Odani, S; Odani, S

    1998-04-01

    A 10-kDa protein was isolated from resting seeds of hemp (Cannabis sativa) by buffer extraction, gel filtration, ion-exchange chromatography, and reversed-phase high-pressure liquid chromatography. The protein did not inhibit bovine trypsin. It consisted of subunits composed of 27 and 61 residues and was held together by two disulfide bonds. The complete amino acid sequence was identified by protein analysis, and had 20 mole% of amino acids containing sulfur. The protein was most similar to a methionine-rich protein of Brazil nut (Bertholletia excelsa) and to Mabinlin IV, a sweetness-inducing protein of Capparis masaikai. The high methionine content and the absence of trypsin inhibitory activity suggested that the seed protein can be used to improve the nutritional quality of plant food-stuffs.

  7. Regulation of thrombosis and vascular function by protein methionine oxidation.

    PubMed

    Gu, Sean X; Stevens, Jeff W; Lentz, Steven R

    2015-06-18

    Redox biology is fundamental to both normal cellular homeostasis and pathological states associated with excessive oxidative stress. Reactive oxygen species function not only as signaling molecules but also as redox regulators of protein function. In the vascular system, redox reactions help regulate key physiologic responses such as cell adhesion, vasoconstriction, platelet aggregation, angiogenesis, inflammatory gene expression, and apoptosis. During pathologic states, altered redox balance can cause vascular cell dysfunction and affect the equilibrium between procoagulant and anticoagulant systems, contributing to thrombotic vascular disease. This review focuses on the emerging role of a specific reversible redox reaction, protein methionine oxidation, in vascular disease and thrombosis. A growing number of cardiovascular and hemostatic proteins are recognized to undergo reversible methionine oxidation, in which methionine residues are posttranslationally oxidized to methionine sulfoxide. Protein methionine oxidation can be reversed by the action of stereospecific enzymes known as methionine sulfoxide reductases. Calcium/calmodulin-dependent protein kinase II is a prototypical methionine redox sensor that responds to changes in the intracellular redox state via reversible oxidation of tandem methionine residues in its regulatory domain. Several other proteins with oxidation-sensitive methionine residues, including apolipoprotein A-I, thrombomodulin, and von Willebrand factor, may contribute to vascular disease and thrombosis.

  8. Ontogenetic changes in digestive enzyme activities and the amino acid profile of starry flounder Platichthys stellatus

    NASA Astrophysics Data System (ADS)

    Song, Zhidong; Wang, Jiying; Qiao, Hongjin; Li, Peiyu; Zhang, Limin; Xia, Bin

    2016-09-01

    Ontogenetic changes in digestive enzyme activities and the amino acid (AA) profile of starry flounder, Platichthys stellatus, were investigated and limiting amino acids were estimated compared with the essential AA profile between larvae and live food to clarify starry flounder larval nutritional requirements. Larvae were collected at the egg stage and 0, 2, 4, 7, 12, 17, 24 days after hatching (DAH) for analysis. Larvae grew from 1.91 mm at hatching to 12.13 mm at 24 DAH. Trypsin and chymotrypsin activities changed slightly by 4 DAH and then increased significantly 4 DAH. Pepsin activity increased sharply beginning 17 DAH. Lipase activity increased significantly 4 DAH and increased progressively with larval growth. Amylase activity was also detected in newly hatched larvae and increased 7 DAH followed by a gradual decrease. High free amino acid (FAA) content was detected in starry flounder eggs (110.72 mg/g dry weight). Total FAA content dropped to 43.29 mg/g in 4-DAH larvae and then decreased gradually to 13.74 mg/g in 24-DAH larvae. Most FAAs (except lysine and methionine) decreased >50% in 4-DAH larvae compared with those in eggs and then decreased to the lowest values in 24-DAH larvae. Changes in the protein amino acid (PAA) profile were much milder than those observed for FAAs. Most PAAs increased gradually during larval development, except lysine and phenylalanine. The percentages of free threonine, valine, isoleucine, and leucine decreased until the end of the trial, whereas the protein forms of these four AAs followed the opposite trend. A comparison of the essential AA composition of live food (rotifers, Artemia nauplii, and Artemia metanauplii) and larvae suggested that methionine was potentially the first limiting AA. These results may help develop starry flounder larviculture methods by solving the AA imbalance in live food. Moreover, the increased digestive enzyme activities indicate the possibility of introducing artificial compound feed.

  9. Differential distribution of amino acids in plants.

    PubMed

    Kumar, Vinod; Sharma, Anket; Kaur, Ravdeep; Thukral, Ashwani Kumar; Bhardwaj, Renu; Ahmad, Parvaiz

    2017-05-01

    Plants are a rich source of amino acids and their individual abundance in plants is of great significance especially in terms of food. Therefore, it is of utmost necessity to create a database of the relative amino acid contents in plants as reported in literature. Since in most of the cases complete analysis of profiles of amino acids in plants was not reported, the units used and the methods applied and the plant parts used were different, amino acid contents were converted into relative units with respect to lysine for statistical analysis. The most abundant amino acids in plants are glutamic acid and aspartic acid. Pearson's correlation analysis among different amino acids showed that there were no negative correlations between the amino acids. Cluster analysis (CA) applied to relative amino acid contents of different families. Alismataceae, Cyperaceae, Capparaceae and Cactaceae families had close proximity with each other on the basis of their relative amino acid contents. First three components of principal component analysis (PCA) explained 79.5% of the total variance. Factor analysis (FA) explained four main underlying factors for amino acid analysis. Factor-1 accounted for 29.4% of the total variance and had maximum loadings on glycine, isoleucine, leucine, threonine and valine. Factor-2 explained 25.8% of the total variance and had maximum loadings on alanine, aspartic acid, serine and tyrosine. 14.2% of the total variance was explained by factor-3 and had maximum loadings on arginine and histidine. Factor-4 accounted 8.3% of the total variance and had maximum loading on the proline amino acid. The relative content of different amino acids presented in this paper is alanine (1.4), arginine (1.8), asparagine (0.7), aspartic acid (2.4), cysteine (0.5), glutamic acid (2.8), glutamine (0.6), glycine (1.0), histidine (0.5), isoleucine (0.9), leucine (1.7), lysine (1.0), methionine (0.4), phenylalanine (0.9), proline (1.1), serine (1.0), threonine (1

  10. Mechanistic and Kinetic Study of Singlet O2 Oxidation of Methionine by On-Line Electrospray Ionization Mass Spectrometry

    NASA Astrophysics Data System (ADS)

    Liu, Fangwei; Lu, Wenchao; Yin, Xunlong; Liu, Jianbo

    2016-01-01

    We report a reaction apparatus developed to monitor singlet oxygen (1O2) reactions in solution using on-line ESI mass spectrometry and spectroscopy measurements. 1O2 was generated in the gas phase by the reaction of H2O2 with Cl2, detected by its emission at 1270 nm, and bubbled into aqueous solution continuously. 1O2 concentrations in solution were linearly related to the emission intensities of airborne 1O2, and their absolute scales were established based on a calibration using 9,10-anthracene dipropionate dianion as an 1O2 trapping agent. Products from 1O2 oxidation were monitored by UV-Vis absorption and positive/negative ESI mass spectra, and product structures were elucidated using collision-induced dissociation-tandem mass spectrometry. To suppress electrical discharge in negative ESI of aqueous solution, methanol was added to electrospray via in-spray solution mixing using theta-glass ESI emitters. Capitalizing on this apparatus, the reaction of 1O2 with methionine was investigated. We have identified methionine oxidation intermediates and products at different pH, and measured reaction rate constants. 1O2 oxidation of methionine is mediated by persulfoxide in both acidic and basic solutions. Persulfoxide continues to react with another methionine, yielding methionine sulfoxide as end-product albeit with a much lower reaction rate in basic solution. Density functional theory was used to explore reaction potential energy surfaces and establish kinetic models, with solvation effects simulated using the polarized continuum model. Combined with our previous study of gas-phase methionine ions with 1O2, evolution of methionine oxidation pathways at different ionization states and in different media is described.

  11. Mechanistic and Kinetic Study of Singlet O2 Oxidation of Methionine by On-Line Electrospray Ionization Mass Spectrometry.

    PubMed

    Liu, Fangwei; Lu, Wenchao; Yin, Xunlong; Liu, Jianbo

    2016-01-01

    We report a reaction apparatus developed to monitor singlet oxygen ((1)O2) reactions in solution using on-line ESI mass spectrometry and spectroscopy measurements. (1)O2 was generated in the gas phase by the reaction of H2O2 with Cl2, detected by its emission at 1270 nm, and bubbled into aqueous solution continuously. (1)O2 concentrations in solution were linearly related to the emission intensities of airborne (1)O2, and their absolute scales were established based on a calibration using 9,10-anthracene dipropionate dianion as an (1)O2 trapping agent. Products from (1)O2 oxidation were monitored by UV-Vis absorption and positive/negative ESI mass spectra, and product structures were elucidated using collision-induced dissociation-tandem mass spectrometry. To suppress electrical discharge in negative ESI of aqueous solution, methanol was added to electrospray via in-spray solution mixing using theta-glass ESI emitters. Capitalizing on this apparatus, the reaction of (1)O2 with methionine was investigated. We have identified methionine oxidation intermediates and products at different pH, and measured reaction rate constants. (1)O2 oxidation of methionine is mediated by persulfoxide in both acidic and basic solutions. Persulfoxide continues to react with another methionine, yielding methionine sulfoxide as end-product albeit with a much lower reaction rate in basic solution. Density functional theory was used to explore reaction potential energy surfaces and establish kinetic models, with solvation effects simulated using the polarized continuum model. Combined with our previous study of gas-phase methionine ions with (1)O2, evolution of methionine oxidation pathways at different ionization states and in different media is described.

  12. Gender differences in methionine accumulation and metabolism in freshly isolated mouse hepatocytes: Potential roles in toxicity

    SciTech Connect

    Dever, Joseph T.; Elfarra, Adnan A.

    2009-05-01

    L-Methionine (Met) is hepatotoxic at high concentrations. Because Met toxicity in freshly isolated mouse hepatocytes is gender-dependent, the goal of this study was to assess the roles of Met accumulation and metabolism in the increased sensitivity of male hepatocytes to Met toxicity compared with female hepatocytes. Male hepatocytes incubated with Met (30 mM) at 37 {sup o}C exhibited higher levels of intracellular Met at 0.5, 1.0, and 1.5 h, respectively, compared to female hepatocytes. Conversely, female hepatocytes had higher levels of S-adenosyl-L-methionine compared to male hepatocytes. Female hepatocytes also exhibited higher L-methionine-L-sulfoxide levels relative to control hepatocytes, whereas the increases in L-methionine-D-sulfoxide (Met-D-O) levels were similar in hepatocytes of both genders. Addition of aminooxyacetic acid (AOAA), an inhibitor of Met transamination, significantly increased Met levels at 1.5 h and increased Met-D-O levels at 1.0 and 1.5 h only in Met-exposed male hepatocytes. No gender differences in cytosolic Met transamination activity by glutamine transaminase K were detected. However, female mouse liver cytosol exhibited higher methionine-DL-sulfoxide (MetO) reductase activity than male mouse liver cytosol at low (0.25 and 0.5 mM) MetO concentrations. Collectively, these results suggest that increased cellular Met accumulation, decreased Met transmethylation, and increased Met and MetO transamination in male mouse hepatocytes may be contributing to the higher sensitivity of the male mouse hepatocytes to Met toxicity in comparison with female mouse hepatocytes.

  13. Gender differences in methionine accumulation and metabolism in freshly isolated mouse hepatocytes: potential roles in toxicity.

    PubMed

    Dever, Joseph T; Elfarra, Adnan A

    2009-05-01

    L-methionine (Met) is hepatotoxic at high concentrations. Because Met toxicity in freshly isolated mouse hepatocytes is gender-dependent, the goal of this study was to assess the roles of Met accumulation and metabolism in the increased sensitivity of male hepatocytes to Met toxicity compared with female hepatocytes. Male hepatocytes incubated with Met (30 mM) at 37 degrees C exhibited higher levels of intracellular Met at 0.5, 1.0, and 1.5 h, respectively, compared to female hepatocytes. Conversely, female hepatocytes had higher levels of S-adenosyl-L-methionine compared to male hepatocytes. Female hepatocytes also exhibited higher L-methionine-L-sulfoxide levels relative to control hepatocytes, whereas the increases in L-methionine-D-sulfoxide (Met-D-O) levels were similar in hepatocytes of both genders. Addition of aminooxyacetic acid (AOAA), an inhibitor of Met transamination, significantly increased Met levels at 1.5 h and increased Met-d-O levels at 1.0 and 1.5 h only in Met-exposed male hepatocytes. No gender differences in cytosolic Met transamination activity by glutamine transaminase K were detected. However, female mouse liver cytosol exhibited higher methionine-dl-sulfoxide (MetO) reductase activity than male mouse liver cytosol at low (0.25 and 0.5 mM) MetO concentrations. Collectively, these results suggest that increased cellular Met accumulation, decreased Met transmethylation, and increased Met and MetO transamination in male mouse hepatocytes may be contributing to the higher sensitivity of the male mouse hepatocytes to Met toxicity in comparison with female mouse hepatocytes.

  14. The distribution of methionine-enkephalin and leucine-enkephalin in the brain and peripheral tissues

    PubMed Central

    Hughes, J.; Kosterlitz, H.W.; Smith, T.W.

    1977-01-01

    1 A method is described for the rapid extraction of opioid peptides from the brain and other tissues. The method is based on acid extraction of tissues followed by adsorption of the extract onto Amberlite XAD-2 resin. Elution with methanol separates the enkephalins and α-endorphin from β-endorphin. 2 Over 90% of the opioid peptide activity isolated from brain and gut of several species by our method was due to methionine- and leucine-enkephalin. In contrast, the major opioid peptide activity recovered from the pituitary was due to peptides of much greater mol. wt. than the enkephalins. 3 An opioid peptide with properties unlike those of the known endorphins or enkephalins was present in brain extracts. This peptide, termed ε-endorphin, has an apparent mol. wt. of 700 to 1200; it constituted between 5 to 10% of the total opioid activity in our extracts. 4 A differential assay of methionine- and leucine-enkephalin was made either by destroying methionine-enkephalin activity with cyanogen bromide or by separating the peptides by thin layer chromatography. 5 The ratio of methionine-enkephalin to leucine-enkephalin varied greatly in different brain regions. The highest proportions of leucine-enkephalin were found in the cerebral cortex and hippocampus. 6 Formaldehyde perfusion and fixation of the brain in vivo had no significant effect on the brain content of enkephalin, indicating that proteolytic breakdown is not a major problem in the extraction of these peptides. 7 It is suggested that the enkephalins may have a neurotransmitter role in both brain and peripheral tissues and that methionine- and leucine-enkephalin may subserve separate neuronal functions. PMID:597668

  15. THE DISTRIBUTION OF METHIONINE-ENKEPHALIN AND LEUCINE-ENKEPHALIN IN THE BRAIN AND PERIPHERAL TISSUES

    PubMed Central

    Hughes, J; Kosterlitz, HW; Smith, TW

    1997-01-01

    A method is described for the rapid extraction of opioid peptides from the brain and other tissues. The method is based on acid extraction of tissues followed by adsorption of the extract onto Amberlite XAD-2 resin. Elution with methanol separates the enkephalins and α-endorphin from β-endorphin. Over 90% of the opioid peptide activity isolated from brain and gut of several species by our method was due to methionine- and leucine-enkephalin. In contrast, the major opioid peptide activity recovered from the pituitary was due to peptides of much greater mol. wt. than the enkephalins. An opioid peptide with properties unlike those of the known endorphins or enkephalins was present in brain extracts. This peptide, termed ∈-endorphin, has an apparent mol. wt. of 700 to 1200; it constituted between 5 to 10% of the total opioid activity in our extracts. A differential assay of methionine- and leucine-enkephalin was made either by destroying methionine-enkephalin activity with cyanogen bromide or by separating the peptides by thin layer chromatography. The ratio of methionine-enkephalin to leucine-enkephalin varied greatly in different brain regions. The highest proportions of leucine-enkephalin were found in the cerebral cortex and hippocampus. Formaldehyde perfusion and fixation of the brain in vivo had no significant effect on the brain content of enkephalin, indicating that proteolytic breakdown is not a major problem in the extraction of these peptides. It is suggested that the enkephalins may have a neurotransmitter role in both brain and peripheral tissues and that methionine- and leucine-enkephalin may subserve separate neuronal functions. PMID:9142421

  16. Metabolically engineered soybean seed with enhanced threonine levels: biochemical characterization and seed-specific expression of lysine-insensitive variants of aspartate kinases from the enteric bacterium Xenorhabdus bovienii.

    PubMed

    Qi, Qungang; Huang, Jintai; Crowley, James; Ruschke, Lisa; Goldman, Barry S; Wen, Li; Rapp, William D

    2011-02-01

    Threonine (Thr) is one of a few limiting essential amino acids (EAAs) in the animal feed industry, and its level in feed rations can impact production of important meat sources, such as swine and poultry. Threonine as well as EAAs lysine (Lys) and methionine (Met) are all synthesized via the aspartate family pathway. Here, we report a successful strategy to produce high free threonine soybean seed via identification of a feedback-resistant aspartate kinase (AK) enzyme that can be over-expressed in developing soybean seed. Towards this goal, we have purified and biochemically characterized AK from the enteric bacterium Xenorhabdus bovienii (Xb). Site-directed mutagenesis of XbAK identified two key regulatory residues Glu-257 and Thr-359 involved in lysine inhibition. Three feedback-resistant alleles, XbAK_T359I, XbAK_E257K and XbAK_E257K/T359I, have been generated. This study is the first to kinetically characterize the XbAK enzyme and provide biochemical and transgenic evidence that Glu-257 near the catalytic site is a critical residue for the allosteric regulation of AK. Furthermore, seed-specific expression of the feedback-resistant XbAK_T359I or XbAK_E257K allele results in increases of free Thr levels of up to 100-fold in R(1) soybean seed when compared to wild-type. Expression of feedback-sensitive wild-type AK did not substantially impact seed Thr content. In addition to high Thr, transgenic seed also showed substantial increases in other major free amino acid (FAA) levels, resulting in an up to 3.5-fold increase in the total FAA content. The transgenic seed was normal in appearance and germinated well under greenhouse conditions.

  17. Lysine acetylation and cancer: A proteomics perspective.

    PubMed

    Gil, Jeovanis; Ramírez-Torres, Alberto; Encarnación-Guevara, Sergio

    2017-01-06

    Lysine acetylation is a reversible modification controlled by two groups of enzymes: lysine acetyltransferases (KATs) and lysine deacetylases (KDACs). Acetylated lysine residues are recognized by bromodomains, a family of evolutionarily conserved domains. The use of high-resolution mass spectrometry-based proteomics, in combination with the enrichment of acetylated peptides through immunoprecipitation with anti-acetyl-lysine antibodies, has expanded the number of acetylated proteins from histones and a few nuclear proteins to more than 2000 human proteins. Because acetylation targets almost all cellular processes, this modification has been associated with cancer. Several KATs, KDACs and bromodomain-containing proteins have been linked to cancer development. Many small molecules targeting some of these proteins have been or are being tested as potential cancer therapies. The stoichiometry of lysine acetylation has not been explored in cancer, representing a promising field in which to increase our knowledge of how this modification is affected in cancer. In this review, we will focus on the strategies that can be used to go deeper in the characterization of the protein lysine acetylation emphasizing in cancer research.

  18. L-methionine degradation potentialities of cheese-ripening microorganisms.

    PubMed

    Bonnarme, P; Lapadatescu, C; Yvon, M; Spinnler, H E

    2001-11-01

    Volatile sulphur compounds are major flavouring compounds in many traditional fermented foods including cheeses. These compounds are products of the catabolism of L-methionine by cheese-ripening microorganisms. The diversity of L-methionine degradation by such microorganisms, however, remains to be characterized. The objective of this work was to compare the capacities to produce volatile sulphur compounds by five yeasts, Geotrichum candidum, Yarrowia lipolytica, Kluyveromyces lactis, Debaryomyces hansenii, Saccharomyces cerevisiae and five bacteria, Brevibacterium linens, Corynebacterium glutamicum, Arthrobacter sp., Micrococcus lutens and Staphylococcus equorum of technological interest for cheese-ripening. The ability of whole cells of these microorganisms to generate volatile sulphur compounds from L-methionine was compared. The microorganisms produced a wide spectrum of sulphur compounds including methanethiol, dimethylsulfide, dimethyldisulfide, dimethyltrisulfide and also S-methylthioesters, which varied in amount and type according to strain. Most of the yeasts produced methanethiol, dimethylsulfide, dimethyldisulfide and dimethyltrisulfide but did not produce S-methylthioesters, apart from G. candidum that produced S-methyl thioacetate. Bacteria, especially Arth. sp. and Brevi. linens, produced the highest amounts and the greatest variety of volatile sulphur compounds includling methanethiol, sulfides and S-methylthioesters, e.g. S-methyl thioacetate, S-methyl thiobutyrate, S-methyl thiopropionate and S-methyl thioisovalerate. Cell-free extracts of all the yeasts and bacteria were examined for the activity of enzymes possibly involved in L-methionine catabolism, i.e. L-methionine demethiolase, L-methionine aminotransferase and L-methionine deaminase. They all possessed L-methionine demethiolase activity, while some (K. lactis, Deb. hansenii, Arth. sp., Staph. equorum) were deficient in L-methionine aminotransferase, and none produced L-methionine deaminase

  19. Characterization of a pyridoxal-5'-phosphate-dependent l-lysine decarboxylase/oxidase from Burkholderia sp. AIU 395.

    PubMed

    Sugawara, Asami; Matsui, Daisuke; Takahashi, Narumi; Yamada, Miwa; Asano, Yasuhisa; Isobe, Kimiyasu

    2014-11-01

    A novel enzyme, which catalyzed decarboxylation of l-lysine into cadaverine with release of carbon dioxide and oxidative deamination of l-lysine into l-2-aminoadipic 5-semialdehyde with release of ammonia and hydrogen peroxide, was found from a newly isolated Burkholderia sp. AIU 395. The enzyme was specific to l-lysine and did not exhibit enzyme activities for other l-amino acids, l-lysine derivatives, d-amino acids, and amines. The apparent Km values for l-lysine in the oxidation and decarboxylation reactions were estimated to be 0.44 mM and 0.84 mM, respectively. The molecular mass was estimated to be 150 kDa, which was composed of two identical subunits with molecular mass of 76.5 kDa. The enzyme contained one mol of pyridoxal 5'-phosphate per subunit as a prosthetic group. The enzyme exhibiting decarboxylase and oxidase activities for l-lysine was first reported here, while the deduced amino acid sequence was homologous to that of putative lysine decarboxylases from the genus Burkholderia.

  20. Serum amino acid concentrations in patients receiving total parenteral nutrition with an amino acid plus dextrose mixture.

    PubMed

    Philcox, J C; Hartley, T F; Worthley, L I; Thomas, D W

    1984-01-01

    The results of monitoring the serum amino acid concentrations during three infusion regimens using a 5:4 mixture of 70% glucose and the synthetic L-amino acid solution, Synthamin 17 (Travasol) are reported. Twelve stabilized patients received continuous total parenteral nutrition (TPN), eight of whom were subsequently placed on a second regimen of cyclical feeding. A separate group of five patients was infused with amino acids, both with and without simultaneous glucose. The serum amino acid concentrations indicated that the supply of valine, leucine, isoleucine, lysine, and histidine, and the synthesis of taurine from the infused methionine was suboptimal, particularly if the period of TPN was prolonged. The synthesis of tyrosine from phenylalanine appeared to be inversely proportional to the infusion rate of the TPN mixture, in particular the glucose component, resulting in depressed tyrosine and increased phenylalanine concentrations in serum during continuous iv nutrition. Cyclical infusions, on the other hand, permitted the tyrosine and phenylalanine concentrations to return to normal during the noninfusion stage of the cycle. Amino acid measurements enabled us to design an amino acids additive mixture which normalized the serum concentrations in three long-term home parenteral nutrition patients. As a result of these investigations serum amino acid measurements are used routinely to monitor the efficacy of TPN and accommodate any specific amino acid requirements of individual patients.

  1. Comparisons between true digestibility of total nitrogen and limiting amino acids in vegetable proteins fed to rats.

    PubMed

    Sarwar, G; Peace, R W

    1986-07-01

    Values (%) for true digestibility (TD) of protein and individual amino acids in some vegetable proteins were determined by the rat balance (fecal) method. Diets containing 8% crude protein (N X 6.25) from soaked and autoclaved samples of Trapper and Century field peas, lentil, pinto bean, seafarer bean, black bean or fababean and autoclaved samples of soybean, peanut, sunflower, rolled oat, rice + soybean and corn + pea were tested in two rat balance studies. In the case of blends, each protein source provided 50% of total protein. The beans, peas and lentil proteins were limiting in sulphur amino acids, tryptophan and threonine, whereas sunflower and rolled oat were most limiting in lysine. In beans, peas and lentil, the TD values of methionine (51-82), cystine (46-85), tryptophan (47-90) and threonine (62-84) were considerably lower than the TD values of total nitrogen (72-90). Similarly, in sunflower and rolled oat, the TD values of lysine (81-83) were lower than the TD values of total nitrogen (90-91). These data suggested that crude protein digestibility may not be a good predictor of bioavailability of limiting amino acids in vegetable proteins. Amino acid scores of the vegetable proteins were 62-96%. The corrections for true digestibility of protein and individual amino acids lowered the scores by 6-15 and 11-47 percentage units, respectively.

  2. Effects of proteome rebalancing and sulfur nutrition on the accumulation of methionine rich d-zein in transgenic soybeans

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Expression of heterologous methionine-rich proteins to increase the overall sulfur amino acid content of soybean seeds has been only marginally successful, presumably due to low accumulation of transgenes in soybeans. Proteome rebalancing of seed proteins has been shown to promote the accumulation o...

  3. Lysine fortification of wheat flour improves selected indices of the nutritional status of predominantly cereal-eating families in Pakistan.

    PubMed

    Hussain, Tajammal; Abbas, Shaid; Khan, Mushtaq A; Scrimshaw, Nevin S

    2004-06-01

    Wheat provides more than 50% of the protein and calorie intake of the population of Pakistan. Legumes and animal protein that could complement the amino acid pattern of wheat, in which lysine is the first limiting amino acid for utilization of protein, are not affordable by members of lower socioeconomic groups in developing countries. The purpose of the study was to determine whether lysine fortification of wheat flour would have a positive impact on populations consuming a predominantly wheat-based diet. A double-blind study was carried out for three months on the outskirts of Peshawar, Pakistan. Forty families received wheat flour fortified with lysine, and 40 families received wheat flour without lysine. Wheat provided 59% of the protein for men, 65% for women, and 58% for children. The weight and height of the children in both groups increased during the study, but the increase was significantly greater in the lysine group. Hemoglobin increased significantly in the women receiving lysine-fortified flour. Transferrin levels increased significantly in men, women, and children in the lysine group as compared with those in the control group. Prealbumin increased significantly in adults receiving additional lysine but decreased in children. Men, women, and children in the lysine-supplemented families had significant increases in CD4, CD8, and complement C3 as compared with controls. These results indicate that lysine fortification of wheat flour can significantly improve sensitive indicators of nutritional status in a population consuming a diet in which 58% to 65% of the protein, depending on age and sex, is supplied by wheat.

  4. Effects of diet supplementation with white tea and methionine on lipid metabolism of gilthead sea bream juveniles (Sparus aurata).

    PubMed

    Pérez-Jiménez, Amalia; Peres, Helena; Rubio, Vera Cruz; Oliva-Teles, Aires

    2013-06-01

    A growth trial was performed with gilthead sea bream juveniles (Sparus aurata) to evaluate the effect of diet supplementation with white tea and methionine on fish performance and lipid metabolism. For that purpose, four diets were formulated: a fish meal-based diet (Control) and diets identical to the control diet but supplemented with 2.9 % white tea (Tea), 0.3 % methionine (Met) or 2.9 % white tea plus 0.3 % methionine (Tea + Met). Growth performance and feed efficiency parameters, whole-body and liver composition, plasma metabolites concentration and liver glucose 6-phosphate dehydrogenase (G6PDH), malic enzyme (ME) and fatty acid synthetase (FAS) activities were determined. Feed intake was higher in fish fed methionine-supplemented diets, whereas this parameter and growth was decreased in fish fed white tea supplementation. Feed efficiency and protein efficiency ratio were not affected by diet composition. Plasma HDL cholesterol and total lipids concentration were higher in fish fed white tea-supplemented diets. Whole-body lipid, plasma glucose, liver glycogen concentration and liver G6PDH, ME and FAS activities were lower in fish fed white tea-supplemented diets. Results of the present study indicate that methionine seems to act as a feed attractant in diets for sea bream juveniles. Additionally, white tea is an important modulator of lipid metabolism in sea bream juveniles.

  5. Proteome-wide enrichment of proteins modified by lysine methylation

    PubMed Central

    Carlson, Scott M; Moore, Kaitlyn E; Green, Erin M; Martín, Glòria Mas; Gozani, Or

    2015-01-01

    We present a protocol for using the triple malignant brain tumor domains of L3MBTL1 (3×MBT), which bind to mono- and di-methylated lysine with minimal sequence specificity, in order to enrich for such methylated lysine from cell lysates. Cells in culture are grown with amino acids containing light or heavy stable isotopic labels. Methylated proteins are enriched by incubating cell lysates with 3×MBT, or with the binding-null D355N mutant as a negative control. Quantitative liquid chromatography and tandem mass spectrometry (LC-MS/MS) are then used to identify proteins that are specifically enriched by 3×MBT pull-down. The addition of a third isotopic label allows the comparison of protein lysine methylation between different biological conditions. Unlike most approaches, our strategy does not require a prior hypothesis of candidate methylated proteins, and it recognizes a wider range of methylated proteins than any available method using antibodies. Cells are prepared by growing in isotopic labeling medium for about 7 d; the process of enriching methylated proteins takes 3 d and analysis by LC-MS/MS takes another 1–2 d. PMID:24309976

  6. Analysis of embryo, cytoplasmic and maternal genetic correlations for seven essential amino acids in rapeseed meal (Brassica napus L.).

    PubMed

    Chen, Guo Lin; Wu, Jian Guo; Variath, Murali-Tottekkaad; Yang, Zhong Wei; Shi, Chun Hai

    2011-04-01

    Genetic correlations of nutrient quality traits including lysine, methionine, leucine, isoleucine, phenylalanine, valine and threonine contents in rapeseed meal were analysed by the genetic model for quantitative traits of diploid plants using a diallel design with nine parents of Brassica napus L. These results indicated that the genetic correlations of embryo, cytoplasm and/or maternal plant havemade different contribution to total genetic correlations of most pairwise nutrient quality traits. The genetic correlations among the amino acids in rapeseed meal were simultaneously controlled by genetic main correlations and genotype x environment (GE) interaction correlations, especially for the maternal dominance correlations. Most components of genetic main correlations and GE interaction correlations for the pairwise traits studied were significantly positive. Some of the pairwise traits had negative genetic correlations, especially between valine and other amino acid contents. Indirect selection for improving the quality traits of rapeseed meal could be expected in rape breeding according to the magnitude and direction of genetic correlation components.

  7. Extending the cross-linking/mass spectrometry strategy: Facile incorporation of photo-activatable amino acids into the model protein calmodulin in Escherichia coli cells.

    PubMed

    Piotrowski, Christine; Ihling, Christian H; Sinz, Andrea

    2015-11-01

    Photo-induced cross-linking is a highly promising technique to investigate protein conformations and protein-protein interactions in their natural cellular environment. One strategy relies on the non-directed incorporation of diazirine-containing photo-activatable amino acids into proteins and a subsequent cross-link formation induced by UV-A irradiation. The advantage of this photo-cross-linking strategy is that it is not restricted to lysine residues and that hydrophobic regions in proteins can also be targeted, which is advantageous for investigating membrane proteins. Here, we present a simplified protocol that relies on the use of mineral salts medium without any special requirements for the incorporation of photo-methionines into proteins in Escherichia coli cells. The possibility to perform these experiments in E. coli is especially valuable as it is the major system for recombinant protein production. The method is exemplified for the Ca(2+) regulating protein calmodulin containing nine methionines, which were found to be replaced by their photo-activatable analogues. Our protocol allows the facile and stochastic incorporation of photo-methionines as the basis for conducting photo-cross-linking experiments in E. coli in an efficient manner.

  8. Mulberry (Morus L.) methionine sulfoxide rreductase gene cloning, sequence analysis, and expression in plant development and stress response.

    PubMed

    Tong, Wei; Zhang, Yinghua; Wang, Heng; Li, Feng; Liu, Zhaoyue; Wang, Yuhua; Fang, Rongjun; Zhao, Weiguo; Li, Long

    2013-01-01

    Methionine sulfoxide reductase plays a regulatory role in plant growth and development, especially in scavenging reactive oxygen species by restoration of the oxidation of methionine in protein. A full-length cDNA sequence encoding methionine sulfoxide reductase (MSR) from mulberry, which we designated MMSR, was cloned based on mulberry expressed sequence tags (ESTs). Sequence analysis showed that the MMSR is 810 bp long, encoding 194 amino acids with a predicted molecular weight of 21.6 kDa and an isoelectric point of 6.78. The expression level of the MMSR gene under conditions of drought and salt stresses was quantified by qRT-PCR. The results show that the expression level changed significantly under the stress conditions compared to the normal growth environment. It helps us to get a better understanding of the molecular basis for signal transduction mechanisms underlying the stress response in mulberry.

  9. Methionine as a Precursor of Ethylene—Commentary

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Lieberman et al. showed in a 1966 publication of Plant Physiology that methionine is a precursor of ethylene. It was the first paper that showed ethylene carbons are derived from carbons 3 and 4 of methionine. This paper catalyzed remarkable interest among plant biologists to elucidate the biosynth...

  10. High Methionine Diet Poses Cardiac Threat: A Molecular Insight.

    PubMed

    Chaturvedi, Pankaj; Kamat, Pradip K; Kalani, Anuradha; Familtseva, Anastasia; Tyagi, Suresh C

    2016-07-01

    High methionine diet (HMD) for example red meat which includes lamb, beef, pork can pose cardiac threat and vascular dysfunction but the mechanisms are unclear. We hypothesize that a diet rich in methionine can malfunction the cardiovascular system in three ways: (1) by augmenting oxidative stress; (2) by inflammatory manifestations; and (3) by matrix/vascular remodeling. To test this hypothesis we used four groups of mice: (1) WT; (2) WT + methionine; (3) CBS(+/-) ; (4) CBS(+/-) +methionine. We observed high oxidative stress in mice fed with methionine which was even higher in CBS(+/-) and CBS(+/-) +methionine. Higher oxidative stress was indicated by high levels of SOD-1 in methionine fed mouse hearts whereas IL-1β, IL-6, TNFα, and TLR4 showed high inflammatory manifestations. The upregulated levels of eNOS/iNOS and upregulated levels of MMP2/MMP9 along with high collagen deposition indicated vascular and matrix remodeling in methionine fed mouse. We evaluated the cardiac function which was dysregulated in the mice fed with HMD. These mice had decreased ejection fraction and left ventricular dysfunction which subsequently leads to adverse cardiac remodeling. In conclusion, our study clearly shows that HMD poses a cardiac threat by increasing oxidative stress, inflammatory manifestations, matrix/vascular remodeling, and decreased cardiac function.

  11. Proteome-wide analysis of lysine acetylation in the plant pathogen Botrytis cinerea

    PubMed Central

    Lv, Binna; Yang, Qianqian; Li, Delong; Liang, Wenxing; Song, Limin

    2016-01-01

    Lysine acetylation is a dynamic and reversible post-translational modification that plays an important role in diverse cellular processes. Botrytis cinerea is the most thoroughly studied necrotrophic species due to its broad host range and huge economic impact. However, to date, little is known about the functions of lysine acetylation in this plant pathogen. In this study, we determined the lysine acetylome of B. cinerea through the combination of affinity enrichment and high-resolution LC-MS/MS analysis. Overall, 1582 lysine acetylation sites in 954 proteins were identified. Bioinformatics analysis shows that the acetylated proteins are involved in diverse biological functions and show multiple cellular localizations. Several particular amino acids preferred near acetylation sites, including KacY, KacH, Kac***R, KacF, FKac and Kac***K, were identified in this organism. Protein interaction network analysis demonstrates that a variety of interactions are modulated by protein acetylation. Interestingly, 6 proteins involved in virulence of B. cinerea, including 3 key components of the high-osmolarity glycerol pathway, were found to be acetylated, suggesting that lysine acetylation plays regulatory roles in pathogenesis. These data provides the first comprehensive view of the acetylome of B. cinerea and serves as a rich resource for functional analysis of lysine acetylation in this plant pathogen. PMID:27381557

  12. Accessibility and mobility of lysine residues in. beta. -lactoglobulin

    SciTech Connect

    Brown, E.M.; Pfeffer, P.E.; Kumosinski, T.F.; Greenberg, R.

    1988-07-26

    N/sup epsilon/-(/sup 2/H/sub 6/)Isopropyllysyl-..beta..-lactoglobulin was prepared by reductive alkylation of ..beta..-lactoglobulin with (/sup 2/H/sub 6/)acetone and NaBH/sub 4/ to provide a /sup 2/H (NMR) probe for the study of lysine involvement in lipid-protein interactions. Amino acid analysis showed 80% of the protein's 15 lysine residues to be labeled. Unmodified lysine residues were located through peptide maps produced from CNBr, tryptic, and chymotryptic digests of the labeled protein. Average correlation times calculated from /sup 2/H NMR spectra were 20 and 320 ps for 8.7 and 3.3 residues, respectively, in 6 M guanidine hydrochloride; in nondenaturing solution, values of 70 and 320 ps were obtained for 6.5 and 3.2 residues, respectively, with the remaining 2.3 modified residues not observed, suggesting that side chains of lysine residues in unordered or flexible regions were more mobile than those in stable periodic structures. /sup 2/H NMR spectra of the protein complexed with dipalmitoylphosphatidylcholine confirmed the extrinsic membrane protein type behavior of ..beta..-lactoglobulin previously reported from /sup 31/P NMR studies of the phospholipids complexed with ..beta..-lactoglobulin. Although no physiological function has yet been identified, comparison of these results with the X-ray structure supports the hypothesis that residues not accessible for modification may help to stabilize the cone-shaped ..beta..-barrel thought to contain binding sites for small lipid-soluble molecules.

  13. Effect of dietary lysine restriction and arginine supplementation in two patients with pyridoxine-dependent epilepsy.

    PubMed

    Yuzyuk, Tatiana; Thomas, Amanda; Viau, Krista; Liu, Aiping; De Biase, Irene; Botto, Lorenzo D; Pasquali, Marzia; Longo, Nicola

    2016-07-01

    Pyridoxine-Dependent Epilepsy (PDE) is a recessive disorder caused by deficiency of α-aminoadipic semialdehyde dehydrogenase in the catabolic pathway of lysine. It is characterized by intractable seizures controlled by the administration of pharmacological doses of vitamin B6. Despite seizure control with pyridoxine, intellectual disability and developmental delays are still observed in some patients with PDE, likely due to the accumulation of toxic intermediates in the lysine catabolic pathway: alpha-aminoadipic semialdehyde (AASA), delta-1-piperideine-6-carboxylate (P6C), and pipecolic acid. Here we evaluate biochemical and clinical parameters in two PDE patients treated with a lysine-restricted diet and arginine supplementation (100-150mg/kg), aimed at reducing the levels of PDE biomarkers. Lysine restriction resulted in decreased accumulation of PDE biomarkers and improved development. Plasma lysine but not plasma arginine, directly correlated with plasma levels of AASA-P6C (p<0.001, r(2)=0.640) and pipecolic acid (p<0.01, r(2)=0.484). In addition, plasma threonine strongly correlated with the levels of AASA-P6C (p<0.0001, r(2)=0.732) and pipecolic acid (p<0.005, r(2)=0.527), suggesting extreme sensitivity of threonine catabolism to pyridoxine availability. Our results further support the use of dietary therapies in combination with pyridoxine for the treatment of PDE.

  14. Nitrogen Sparing Induced by a Mixture of Essential Amino Acids Given Chiefly as Their Keto-Analogues during Prolonged Starvation in Obese Subjects

    PubMed Central

    Sapir, Daniel G.; Owen, Oliver E.; Pozefsky, Thomas; Walser, Mackenzie

    1974-01-01

    11 normal obese subjects were fasted for 33 days. In five, who served as controls, urine urea nitrogen excretion remained constant for 2 wk thereafter. The other six were given seven daily infusions containing 6-8 mmol each of the α-keto-analogues of valine, leucine, isoleucine, phenylalanine, and methionine (as sodium salts) plus 3-4 mmol each of the remaining essential amino acids (lysine, threonine, tryptophan, and histidine). Rapid amination of the infused ketoacids occurred, as indicated by significant increases in plasma concentrations of valine, leucine, isoleucine, alloisoleucine, phenylalanine, and methionine. Glutamine, glycine, serine, glutamate, and taurine fell significantly. Blood glucose, ketone bodies, plasma free fatty acids, and serum immunoreactive insulin concentrations were unaltered. Urine urea nitrogen fell from 1.46 to 0.89 g/day on the last day of infusions; 5 days later it was still lower (0.63 g/day) and in two subjects studied for 9 and 17 days postinfusion it remained below preinfusion control values. Urine ammonia, creatinine, and uric acid were unaltered. Nitrogen balance became less negative during and after infusions. The results indicate that this mixture of essential amino acids and their keto-analogues facilitates nitrogen sparing during prolonged starvation, in part by conversion of the ketoacids to amino acids and in part by altering mechanisms of nitrogen conservation. The latter effect persists after the ketoacids are metabolized. PMID:4430727

  15. Co-expression of bacterial aspartate kinase and adenylylsulfate reductase genes substantially increases sulfur amino acid levels in transgenic alfalfa (Medicago sativa L.).

    PubMed

    Tong, Zongyong; Xie, Can; Ma, Lei; Liu, Liping; Jin, Yongsheng; Dong, Jiangli; Wang, Tao

    2014-01-01

    Alfalfa (Medicago sativa L.) is one of the most important forage crops used to feed livestock, such as cattle and sheep, and the sulfur amino acid (SAA) content of alfalfa is used as an index of its nutritional value. Aspartate kinase (AK) catalyzes the phosphorylation of aspartate to Asp-phosphate, the first step in the aspartate family biosynthesis pathway, and adenylylsulfate reductase (APR) catalyzes the conversion of activated sulfate to sulfite, providing reduced sulfur for the synthesis of cysteine, methionine, and other essential metabolites and secondary compounds. To reduce the feedback inhibition of other metabolites, we cloned bacterial AK and APR genes, modified AK, and introduced them into alfalfa. Compared to the wild-type alfalfa, the content of cysteine increased by 30% and that of methionine increased substantially by 60%. In addition, a substantial increase in the abundance of essential amino acids (EAAs), such as aspartate and lysine, was found. The results also indicated a close connection between amino acid metabolism and the tricarboxylic acid (TCA) cycle. The total amino acid content and the forage biomass tested showed no significant changes in the transgenic plants. This approach provides a new method for increasing SAAs and allows for the development of new genetically modified crops with enhanced nutritional value.

  16. Effect of supplementation of crystalline lysine on the performance of WL layers in tropics during summer.

    PubMed

    Kumari, K Naga Raja; Reddy, V Ravinder; Preetham, V Chinni; Kumar, D Srinivas; Sen, Arup Ratan; Rao, S Venkata Rama

    2016-04-01

    A trial was conducted to evaluate the effect of lysine concentration in the diet of WL layers with constant ratio of other essential amino acids to lysine. Pullets (528) aged 25 to 36 weeks were fed with test diet containing two protein levels (13.36 and 15.78%) each with 5% concentration of lysine (0.50, 0.55, 0.60, 0.65, and 0.70) and a control with 17% CP and 0.70%, lysine. Each test diet was fed ad libitum to six replicates of eight birds for a period of 12 weeks. Egg production (EP), egg weight (EW), egg mass (EM), feed efficiency (g/g) (FE), body weight gain (BWG), Haugh unit (HU) and yolk colour (YC) were measured. Increased (P ≤ 0.05) EP, EW, EM, FE and BWG were obtained with increasing lysine concentration in diets. Whereas, feed intake/h/day, feed intake/egg, egg shell defects (ESD), mortality and shell thickness were not affected (P ≥ 0.05) by the concentration of lysine in diet. However, higher (P ≤ 0.05) HU score and YC were noticed at low lysine (0.50 %) concentrations. Based on this, it was concluded that WL layers (25-36 weeks) reared in open-sided houses in the tropics require approximately 0.70 % lysine (597.90 vs. 584.39 mg/h/day) in low (13.36% CP) and high (15.78% CP) protein groups in diets containing approximately 2700 kcal of ME/kg in summer.

  17. Detection and Measurement of Methionine Oxidation in Proteins.

    PubMed

    Sen, K Ilker; Hepler, Robert; Nanda, Hirsh

    2017-02-02

    Methionine oxidation is a prevalent modification found in proteins both in biological settings and in the manufacturing of biotherapeutic molecules. In cells, the oxidation of specific methionine sites can modulate protein function or promote interactions that trigger signaling pathways. In biotherapeutic development, the formation of oxidative species could be detrimental to the efficacy or safety of the drug product. Thus, methionine oxidation is a critical quality attribute that needs to be monitored throughout development. Here we describe a method using LC/MS/MS to identify site-specific methionine modifications in proteins. Antibodies are stressed with hydrogen peroxide, and the level of Met oxidation is compared to that of reference molecules. The protocols presented here are not specific to methionine and can be used more generally to identify other PTM risk sites in molecules after various types of treatments. © 2017 by John Wiley & Sons, Inc.

  18. Histone lysine methylation and chromatin replication.

    PubMed

    Rivera, Carlos; Gurard-Levin, Zachary A; Almouzni, Geneviève; Loyola, Alejandra

    2014-12-01

    In eukaryotic organisms, the replication of the DNA sequence and its organization into chromatin are critical to maintain genome integrity. Chromatin components, such as histone variants and histone post-translational modifications, along with the higher-order chromatin structure, impact several DNA metabolic processes, including replication, transcription, and repair. In this review we focus on lysine methylation and the relationships between this histone mark and chromatin replication. We first describe studies implicating lysine methylation in regulating early steps in the replication process. We then discuss chromatin reassembly following replication fork passage, where the incorporation of a combination of newly synthesized histones and parental histones can impact the inheritance of lysine methylation marks on the daughter strands. Finally, we elaborate on how the inheritance of lysine methylation can impact maintenance of the chromatin landscape, using heterochromatin as a model chromatin domain, and we discuss the potential mechanisms involved in this process.

  19. SPOTing Acetyl-Lysine Dependent Interactions

    PubMed Central

    Picaud, Sarah; Filippakopoulos, Panagis

    2015-01-01

    Post translational modifications have been recognized as chemical signals that create docking sites for evolutionary conserved effector modules, allowing for signal integration within large networks of interactions. Lysine acetylation in particular has attracted attention as a regulatory modification, affecting chromatin structure and linking to transcriptional activation. Advances in peptide array technologies have facilitated the study of acetyl-lysine-containing linear motifs interacting with the evolutionary conserved bromodomain module, which specifically recognizes and binds to acetylated sequences in histones and other proteins. Here we summarize recent work employing SPOT peptide technology to identify acetyl-lysine dependent interactions and document the protocols adapted in our lab, as well as our efforts to characterize such bromodomain-histone interactions. Our results highlight the versatility of SPOT methods and establish an affordable tool for rapid access to potential protein/modified-peptide interactions involving lysine acetylation. PMID:27600229

  20. A Method to determine lysine acetylation stoichiometries

    SciTech Connect

    Nakayasu, Ernesto S.; Wu, Si; Sydor, Michael A.; Shukla, Anil K.; Weitz, Karl K.; Moore, Ronald J.; Hixson, Kim K.; Kim, Jong Seo; Petyuk, Vladislav A.; Monroe, Matthew E.; Pasa-Tolic, Ljiljana; Qian, Weijun; Smith, Richard D.; Adkins, Joshua N.; Ansong, Charles

    2014-07-21

    A major bottleneck to fully understanding the functional aspects of lysine acetylation is the lack of stoichiometry information. Here we describe a mass spectrometry method using a combination of isotope labeling and detection of a diagnostic fragment ion to determine the stoichiometry of lysine acetylation on proteins globally. Using this technique, we determined the modification occupancy on hundreds of acetylated peptides from cell lysates and cross-validated the measurements via immunoblotting.

  1. Potent and sustained cellular inhibition of miR-122 by lysine-derivatized peptide nucleic acids (PNA) and phosphorothioate locked nucleic acid (LNA)/2'-O-methyl (OMe) mixmer anti-miRs in the absence of transfection agents

    PubMed Central

    Torres, Adrian G.; Threlfall, Richard N.

    2011-01-01

    Efficient cell delivery of antisense oligonucleotides (ONs) is a key issue for their potential therapeutic use. It has been shown recently that some ONs can be delivered into cells without the use of transfection agents (gymnosis), but this generally requires cell incubation over several days and high amounts of ONs (micromolar concentrations). Here we have targeted microRNA 122 (miR-122), a small non-coding RNA involved in regulation of lipid metabolism and in the replication of hepatitis C virus, with ONs of different chemistries (anti-miRs) by gymnotic delivery in cell culture. Using a sensitive dual-luciferase reporter assay, anti-miRs were screened for their ability to enter liver cells gymnotically and inhibit miR-122 activity. Efficient miR-122 inhibition was obtained with cationic PNAs and 2'-O-methyl (OMe) and Locked Nucleic Acids (LNA)/OMe mixmers containing either phosphodiester (PO) or phosphorothioate (PS) linkages at sub-micromolar concentrations when incubated with cells for just 4 hours. Furthermore, PNA and PS-containing anti-miRs were able to sustain miR-122 inhibitory effects for at least 4 days. LNA/OMe PS anti-miRs were the most potent anti-miR chemistry tested in this study, an ON chemistry that has been little exploited so far as anti-miR agents towards therapeutics. PMID:22567190

  2. Stabilization of methionine-rich protein in Saccharomyces cerevisiae: targeting of BZN protein into the peroxisome.

    PubMed

    Nicaud, J M; Raynal, A; Beyou, A; Merkamm, M; Ito, H; Labat, N

    1994-01-01

    We have constructed a gene coding for the 12-kDa intermediate form of the 2s methionine-rich protein from Bertholletia excelsa seeds. This protein, expressed intracellularly in yeast, is characterised by a 20-min half-life. By adding 11 amino acids corresponding to the peroxisome-targeting sequence (PTSc) of luciferase, we have significantly increased its half-life. This stabilization allowed accumulation of the BZN protein into the peroxisome as judged by cell fractionation. Accumulation of the 12-kDa protein results in a significant increase of the total methionine content in yeast cells (30%) indicating that such a microorganism could represent a practicable protected shuttle for an animal-feed additive.

  3. Novel fatty acid acylation of lens integral membrane protein aquaporin-0.

    PubMed

    Schey, Kevin L; Gutierrez, Danielle B; Wang, Zhen; Wei, Junhua; Grey, Angus C

    2010-11-16

    Fatty acid acylation of proteins is a well-studied co- or posttranslational modification typically conferring membrane trafficking signals or membrane anchoring properties to proteins. Commonly observed examples of protein acylation include N-terminal myristoylation and palmitoylation of cysteine residues. In the present study, direct tissue profiling mass spectrometry of bovine and human lens sections revealed an abundant signal tentatively assigned as a lipid-modified form of aquaporin-0. LC/MS/MS proteomic analysis of hydrophobic tryptic peptides from lens membrane proteins revealed both N-terminal and C-terminal peptides modified by 238 and 264 Da which were subsequently assigned by accurate mass measurement as palmitoylation and oleoylation, respectively. Specific sites of modification were the N-terminal methionine residue and lysine 238 revealing, for the first time, an oleic acid modification via an amide linkage to a lysine residue. The specific fatty acids involved reflect their abundance in the lens fiber cell plasma membrane. Imaging mass spectrometry indicated abundant acylated AQP0 in the inner cortical region of both bovine and human lenses and acylated truncation products in the lens nucleus. Additional analyses revealed that the lipid-modified forms partitioned exclusively to a detergent-resistant membrane fraction, suggesting a role in membrane domain targeting.

  4. Free amino acid composition in hemolymph and muscle of the ghost crab, Ocypode platytarsis.

    PubMed

    Sankar, R Siva; Yogamoorthi, A

    2012-05-15

    Free amino acid plays an important role in physiological functions. The ghost crab, Ocypode platytarsis caught off the Puducherry sandy beaches, south east coast of India were investigated for the pattern of distribution of free amino acids. The hemolymph and muscle were sampled from male and female crabs and then subjected to free amino acid analysis quantitatively by HPLC. Fourteen free amino acids have been determined in hemolymph and muscle of both sexes in Ocypode platytarsis. The essential amino acids were leucine, lysine, methionine, phenylalanine, threonine, valine and histidine and non-essential amino acids were alanine, arginine, glutamic acid, glycine, serine and tyrosine. The total concentration of free amino acid found in the male and female hemolymph was lower than that in the muscle. Glutamic acid is of higher concentration in the hemolymph whereas histidine found was higher in the muscle of both sexes. The male crab contains a strikingly higher concentration of histidine of about eight times than that of female in muscles. The study revealed that distribution pattern of free amino acids differ in relation to hemolymph and muscle of male and female crabs.

  5. Primary Action of Indole-3-acetic Acid in Crown Gall Tumors

    PubMed Central

    Rausch, Thomas; Kahl, Günter; Hilgenberg, Willy

    1984-01-01

    Exogenously added indole-3-acetic acid at a concentration of 100 micromolars stimulates d-glucose uptake (or 3-O-methyl-d-glucose uptake) by 25% in crown gall tumors induced on potato tuber tissue by Agrobacterium tumefaciens strain C 58. The titration of the endogenous IAA with the auxin antagonist 2-naphthaleneacetic acid at 100 micromolars reduces d-glucose uptake by about 80%. The apparent inhibition constant Ki is 21 micromolars. Other auxin antagonists like 1-naphthoxyacetic acid and 2-(p-chlorophenoxy)-2-methylpropionic acid show similar effects. The uptake of the amino acids leucine, methionine, tryptophan, lysine, and aspartic acid is also inhibited by 2-naphthaleneacetic acid to similar degrees. The auxins 1-naphthaleneacetic acid and 2-naphthoxyacetic acid at concentrations between 10 and 100 micromolars inhibit solute uptake only slightly (inhibition less than 20%). The impact of the results on the postulated role of indole-3-acetic acid as a modifier of the electrochemical proton gradient across the plasmalemma in crown gall tumor tissue is discussed. PMID:16663625

  6. Primary Action of Indole-3-acetic Acid in Crown Gall Tumors: Increase of Solute Uptake.

    PubMed

    Rausch, T; Kahl, G; Hilgenberg, W

    1984-06-01

    Exogenously added indole-3-acetic acid at a concentration of 100 micromolars stimulates d-glucose uptake (or 3-O-methyl-d-glucose uptake) by 25% in crown gall tumors induced on potato tuber tissue by Agrobacterium tumefaciens strain C 58. The titration of the endogenous IAA with the auxin antagonist 2-naphthaleneacetic acid at 100 micromolars reduces d-glucose uptake by about 80%. The apparent inhibition constant K(i) is 21 micromolars. Other auxin antagonists like 1-naphthoxyacetic acid and 2-(p-chlorophenoxy)-2-methylpropionic acid show similar effects. The uptake of the amino acids leucine, methionine, tryptophan, lysine, and aspartic acid is also inhibited by 2-naphthaleneacetic acid to similar degrees. The auxins 1-naphthaleneacetic acid and 2-naphthoxyacetic acid at concentrations between 10 and 100 micromolars inhibit solute uptake only slightly (inhibition less than 20%). The impact of the results on the postulated role of indole-3-acetic acid as a modifier of the electrochemical proton gradient across the plasmalemma in crown gall tumor tissue is discussed.

  7. Gene cloning, recombinant expression, purification and characterization of l-methionine decarboxylase from Streptomyces sp. 590.

    PubMed

    Hayashi, Masaya; Okada, Akane; Yamamoto, Kumiko; Okugochi, Tomomi; Kusaka, Chika; Kudou, Daizou; Nemoto, Michiko; Inagaki, Junko; Hirose, Yuu; Okajima, Toshihide; Tamura, Takashi; Soda, Kenji; Inagaki, Kenji

    2016-12-21

    l-Methionine decarboxylase (MetDC) from Streptomyces sp. 590 depends on pyridoxal 5'-phosphate and catalyzes the non-oxidative decarboxylation of l-methionine to produce 3-methylthiopropylamine and carbon dioxide. MetDC gene (mdc) was determined to consist of 1,674 bp encoding 557 amino acids, and the amino acid sequence is similar to that of l-histidine decarboxylases and l-valine decarboxylases from Streptomyces sp. strains. The mdc gene was cloned and recombinant MetDC was heterologously expressed by Escherichia coli The purification of recombinant MetDC was carried out by DEAE-Toyopearl and Ni-NTA agarose column chromatography. The recombinant enzyme was homodimeric with a molecular mass of 61,000 Da and showed optimal activity between 45 to 55 °C and at pH 6.6, and the stability below 30 °C and between pH 4.6 to 7.0. l-Methionine and l-norleucine were good substrates for MetDC. The Michaelis constants for l-methionine and l-norleucine were 30 and 73 mM, respectively. The recombinant MetDC (0.50 U/ml) severely inhibited growth of human tumour cells A431 (epidermoid ovarian carcinoma cell line) and MDA-MB-231 (breast cancer cell line), however showed relatively low cytotoxicity for human normal cell NHDF-Neo (dermal fibroblast cell line from neonatal foreskin). This study revealed the properties of the gene and the protein sequence of MetDC for the first time.

  8. Metabolic Regulation by Lysine Malonylation, Succinylation, and Glutarylation*

    PubMed Central

    Hirschey, Matthew D.; Zhao, Yingming

    2015-01-01

    Protein acetylation is a well-studied regulatory mechanism for several cellular processes, ranging from gene expression to metabolism. Recent discoveries of new post-translational modifications, including malonylation, succinylation, and glutarylation, have expanded our understanding of the types of modifications found on proteins. These three acidic lysine modifications are structurally similar but have the potential to regulate different proteins in different pathways. The deacylase sirtuin 5 (SIRT5) catalyzes the removal of these modifications from a wide range of proteins in different subcellular compartments. Here, we review these new modifications, their regulation by SIRT5, and their emerging role in cellular regulation and diseases. PMID:25717114

  9. Synthesis, pharmacokinetics, and biological use of lysine-modified single-walled carbon nanotubes

    PubMed Central

    Mulvey, J Justin; Feinberg, Evan N; Alidori, Simone; McDevitt, Michael R; Heller, Daniel A; Scheinberg, David A

    2014-01-01

    We aimed to create a more robust and more accessible standard for amine-modifying single-walled carbon nanotubes (SWCNTs). A 1,3-cycloaddition was developed using an azomethine ylide, generated by reacting paraformaldehyde and a side-chain-Boc (tert-Butyloxycarbonyl)-protected, lysine-derived alpha-amino acid, H-Lys(Boc)-OH, with purified SWCNT or C60. This cycloaddition and its lysine adduct provides the benefits of dense, covalent modification, ease of purification, commercial availability of reagents, and pH-dependent solubility of the product. Subsequently, SWCNTs functionalized with lysine amine handles were covalently conjugated to a radiometalated chelator, 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid (DOTA). The 111In-labeled construct showed rapid renal clearance in a murine model and a favorable biodistribution, permitting utility in biomedical applications. Functionalized SWCNTs strongly wrapped small interfering RNA (siRNA). In the first disclosed deployment of thermophoresis with carbon nanotubes, the lysine-modified tubes showed a desirable, weak SWCNT-albumin binding constant. Thus, lysine-modified nanotubes are a favorable candidate for medicinal work. PMID:25228803

  10. Synthesis, pharmacokinetics, and biological use of lysine-modified single-walled carbon nanotubes.

    PubMed

    Mulvey, J Justin; Feinberg, Evan N; Alidori, Simone; McDevitt, Michael R; Heller, Daniel A; Scheinberg, David A

    2014-01-01

    We aimed to create a more robust and more accessible standard for amine-modifying single-walled carbon nanotubes (SWCNTs). A 1,3-cycloaddition was developed using an azomethine ylide, generated by reacting paraformaldehyde and a side-chain-Boc (tert-Butyloxycarbonyl)-protected, lysine-derived alpha-amino acid, H-Lys(Boc)-OH, with purified SWCNT or C60. This cycloaddition and its lysine adduct provides the benefits of dense, covalent modification, ease of purification, commercial availability of reagents, and pH-dependent solubility of the product. Subsequently, SWCNTs functionalized with lysine amine handles were covalently conjugated to a radiometalated chelator, 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid (DOTA). The (111)In-labeled construct showed rapid renal clearance in a murine model and a favorable biodistribution, permitting utility in biomedical applications. Functionalized SWCNTs strongly wrapped small interfering RNA (siRNA). In the first disclosed deployment of thermophoresis with carbon nanotubes, the lysine-modified tubes showed a desirable, weak SWCNT-albumin binding constant. Thus, lysine-modified nanotubes are a favorable candidate for medicinal work.

  11. 1- sup 13 C; methyl-2H3 methionine kinetics in humans: Methionine conservation and cystine sparing

    SciTech Connect

    Storch, K.J.; Wagner, D.A.; Burke, J.F.; Young, V.R. )

    1990-05-01

    Methionine (Met) conservation in healthy young adult men (4/diet group) was explored by supplying one of the following three L-amino acid based diets: (1) adequate Met but no cystine; (2) neither Met nor cystine; or (3) no Met but cystine supplementation. After 5 days, subjects received a continuous intravenous infusion of L-(1-13C; methyl-2H3)Met for 5 h while the diet was given as small isocaloric isonitrogenous meals. Estimates were made of rates of Met incorporation into protein synthesis (S) and release from body proteins (B), transmethylation (TM), remethylation of homocysteine (RM), and transsulfuration (TS). For the adequate Met diet, the rates were S = 24 +/- 2, B = 18 +/- 1, TM = 12.4 +/- 1.7, RM = 4.7 +/- 1.1, and TS = 7.6 +/- 0.6 (SE) mumol.kg-1.h-1. The sulfur amino acid-devoid diet significantly (P less than 0.05) reduced S, TM, RM, and TS. Supplementation of this diet with cystine reduced Met oxidation (P = 0.05). Therefore, two loci are quantitatively important regulatory points in Met conservation in vivo: (1) the distribution of Met between the pathways of protein anabolism and TM (Met locus) and (2) the distribution of homocysteine between RM and TS (homocysteine locus).

  12. Inversion of the stereochemistry around the sulfur atom of the axial methionine side chain through alteration of amino acid side chain packing in Hydrogenobacter thermophilus cytochrome C552 and its functional consequences.

    PubMed

    Tai, Hulin; Tonegawa, Ken; Shibata, Tomokazu; Hemmi, Hikaru; Kobayashi, Nagao; Yamamoto, Yasuhiko

    2013-07-16

    In cytochrome c, the coordination of the axial Met Sδ atom to the heme Fe atom occurs in one of two distinctly different stereochemical manners, i.e., R and S configurations, depending upon which of the two lone pairs of the Sδ atom is involved in the bond; hence, the Fe-coordinated Sδ atom becomes a chiral center. In this study, we demonstrated that an alteration of amino acid side chain packing induced by the mutation of a single amino acid residue, i.e., the A73V mutation, in Hydrogenobacter thermophilus cytochrome c552 (HT) forces the inversion of the stereochemistry around the Sδ atom from the R configuration [Travaglini-Allocatelli, C., et al. (2005) J. Biol. Chem. 280, 25729-25734] to the S configuration. Functional comparison between the wild-type HT and the A73V mutant possessing the R and S configurations as to the stereochemistry around the Sδ atom, respectively, demonstrated that the redox potential (Em) of the mutant at pH 6.00 and 25 °C exhibited a positive shift of ∼20 mV relative to that of the wild-type HT, i.e., 245 mV, in an entropic manner. Because these two proteins have similar enthalpically stabilizing interactions, the difference in the entropic contribution to the Em value between them is likely to be due to the effect of the conformational alteration of the axial Met side chain associated with the inversion of the stereochemistry around the Sδ atom due to the effect of mutation on the internal mobility of the loop bearing the axial Met. Thus, the present study demonstrated that the internal mobility of the loop bearing the axial Met, relevant to entropic control of the redox function of the protein, is affected quite sensitively by the contextual stereochemical packing of amino acid side chains in the proximity of the axial Met.

  13. Child Stunting is Associated with Low Circulating Essential Amino Acids

    PubMed Central

    Semba, Richard D.; Shardell, Michelle; Sakr Ashour, Fayrouz A.; Moaddel, Ruin; Trehan, Indi; Maleta, Kenneth M.; Ordiz, M. Isabel; Kraemer, Klaus; Khadeer, Mohammed A.; Ferrucci, Luigi; Manary, Mark J.

    2016-01-01

    Background Stunting affects about one-quarter of children under five worldwide. The pathogenesis of stunting is poorly understood. Nutritional interventions have had only modest effects in reducing stunting. We hypothesized that insufficiency in essential amino acids may be limiting the linear growth of children. Methods We used a targeted metabolomics approach to measure serum amino acids, glycerophospholipids, sphingolipids, and other metabolites using liquid chromatography-tandem mass spectrometry in 313 children, aged 12–59 months, from rural Malawi. Children underwent anthropometry. Findings Sixty-two percent of the children were stunted. Children with stunting had lower serum concentrations of all nine essential amino acids (tryptophan, isoleucine, leucine, valine, methionine, threonine, histidine, phenylalanine, lysine) compared with nonstunted children (p < 0.01). In addition, stunted children had significantly lower serum concentrations of conditionally essential amino acids (arginine, glycine, glutamine), non-essential amino acids (asparagine, glutamate, serine), and six different sphingolipids compared with nonstunted children. Stunting was also associated with alterations in serum glycerophospholipid concentrations. Interpretation Our findings support the idea that children with a high risk of stunting may not be receiving an adequate dietary intake of essential amino acids and choline, an essential nutrient for the synthesis of sphingolipids and glycerophospholipids. PMID:27211567

  14. L-lysine as adjunctive treatment in patients with schizophrenia: a single-blinded, randomized, cross-over pilot study

    PubMed Central

    2011-01-01

    Background Accumulating evidence suggests that the brain's nitric oxide (NO) signalling system may be involved in the pathophysiology of schizophrenia and could thus constitute a novel treatment target. The study was designed to investigate the benefit of L-lysine, an amino acid that interferes with NO production, as an add-on treatment for schizophrenia. Methods L-lysine, 6 g/day, was administered to 10 patients with schizophrenia as an adjunctive to their conventional antipsychotic medication. The study was designed as a single-blinded, cross-over study where patients were randomly assigned to initial treatment with either L-lysine or placebo and screened at baseline, after four weeks when treatment was crossed over, and after eight weeks. Results L-lysine treatment caused a significant increase in blood concentration of L-lysine and was well tolerated. A significant decrease in positive symptom severity, measured by the Positive And Negative Syndrome Scale (PANSS), was detected. A certain decrease in score was also observed during placebo treatment and the effects on PANSS could not unequivocally be assigned to the L-lysine treatment. Furthermore, performance on the Wisconsin Card Sorting Test was significantly improved compared to baseline, an effect probably biased by training. Subjective reports from three of the patients indicated decreased symptom severity and enhanced cognitive functioning. Conclusions Four-week L-lysine treatment of 6 g/day caused a significant increase in blood concentration of L-lysine that was well tolerated. Patients showed a significant decrease in positive symptoms as assessed by PANSS in addition to self-reported symptom improvement by three patients. The NO-signalling pathway is an interesting, potentially new treatment target for schizophrenia; however, the effects of L-lysine need further evaluation to decide the amino acid's potentially beneficial effects on symptom severity in schizophrenia. Trial registration NCT00996242 PMID

  15. Simultaneous analysis of Nε-(carboxymethyl)lysine, reducing sugars, and lysine during the dairy thermal process.

    PubMed

    Xu, Xian-Bing; Ma, Fei; Yu, Shu-Juan; Guan, Yong-Guang

    2013-09-01

    A new analytical method allowing the simultaneous quantification of Nε-(carboxymethyl)lysine (CML), lysine, and reducing sugars (glucose, lactose, and galactose) is described. It is based on high performance anion-exchange chromatography with pulsed amperometric electrochemical detection. This method demonstrated a low limit of quantification (0.385 to 0.866 mg/L), excellent linear correlation (R(2)>0.997), and desired calibration range (3.125 to 25 mg/L). In addition, lactose-lysine solutions containing sulfite (4 to 400 mmol/L) were heated at 110°C for 2h. The results showed that sulfite inhibited the formation of CML and promoted the consumption of reducing sugars and lysine in the Maillard reaction model. The method proved to be useful for simultaneous analysis of CML, lysine, and reducing sugars (glucose, galactose, and lactose) in the Maillard reaction system. Moreover, sulfite was an effective inhibitor of CML formation.

  16. Amino acid concentrations in plasma and erythrocytes in aregeneratory and haemolytic anaemias.

    PubMed

    Seip, M; Lindemann, R; Gjesdahl, P; Gjessing, L R

    1975-10-01

    The concentrations of unbound amino acids in erythrocytes and in plasma from 7 normal individuals, 11 patients with various types of aregeneratory anaemia, and 4 patients with hereditary haemolytic anaemias were determined on a Technicon Amino Acid Analyzer (Perry et al 1970). Most amino acids were normally found in higher concentrations in plasma than intracellularly. Cystine, methionine and trypotophan were almost exclusively present in plasma. Aspartic acid, however, was mainly found in erythrocytes, and glutathione only in erythrocytes. Glutamic acid and ornithine were more concentrated in the cells, while glycine and asparagine showed approximately the same concentrations in erythrocytes as in plasma. In the patients, plasma amino acids showed little deviations from normal, but in the erythrocytes there were striking changes. Erythrocyte glutamic acid concentrations were moderately to markedly elevated in all patients studied, and glycine concentrations in 13 out of 15 patients. In addition, the following amino acids were increased intracellularly in more than one patient: glutamine (8 patients), serine (7), asparagine (5), threonine (4), taurine (3), alanine (2), valine (2), ornithine (2), lysine (2), citrulline (2). Aspartic acid was decreased in erythrocytes from 4 patients with aregeneratory and 1 with haemolytic anaemia.

  17. Crystal structure of LL-diaminopimelate aminotransferase from Arabidopsis thaliana: a recently discovered enzyme in the biosynthesis of L-lysine by plants and Chlamydia.

    PubMed

    Watanabe, Nobuhiko; Cherney, Maia M; van Belkum, Marco J; Marcus, Sandra L; Flegel, Mitchel D; Clay, Matthew D; Deyholos, Michael K; Vederas, John C; James, Michael N G

    2007-08-17

    The essential biosynthetic pathway to l-Lysine in bacteria and plants is an attractive target for the development of new antibiotics or herbicides because it is absent in humans, who must acquire this amino acid in their diet. Plants use a shortcut of a bacterial pathway to l-Lysine in which the pyridoxal-5'-phosphate (PLP)-dependent enzyme ll-diaminopimelate aminotransferase (LL-DAP-AT) transforms l-tetrahydrodipicolinic acid (L-THDP) directly to LL-DAP. In addition, LL-DAP-AT was recently found in Chlamydia sp., suggesting that inhibitors of this enzyme may also be effective against such organisms. In order to understand the mechanism of this enzyme and to assist in the design of inhibitors, the three-dimensional crystal structure of LL-DAP-AT was determined at 1.95 A resolution. The cDNA sequence of LL-DAP-AT from Arabidopsis thaliana (AtDAP-AT) was optimized for expression in bacteria and cloned in Escherichia coli without its leader sequence but with a C-terminal hexahistidine affinity tag to aid protein purification. The structure of AtDAP-AT was determined using the multiple-wavelength anomalous dispersion (MAD) method with a seleno-methionine derivative. AtDAP-AT is active as a homodimer with each subunit having PLP in the active site. It belongs to the family of type I fold PLP-dependent enzymes. Comparison of the active site residues of AtDAP-AT and aspartate aminotransferases revealed that the PLP binding residues in AtDAP-AT are well conserved in both enzymes. However, Glu97* and Asn309* in the active site of AtDAP-AT are not found at similar positions in aspartate aminotransferases, suggesting that specific substrate recognition may require these residues from the other monomer. A malate-bound structure of AtDAP-AT allowed LL-DAP and L-glutamate to be modelled into the active site. These initial three-dimensional structures of LL-DAP-AT provide insight into its substrate specificity and catalytic mechanism.

  18. Crystal Structure of Ll-Diaminopimelate Aminotransferase From 'Arabidopsis Thaliana': a Recently-Discovered Enzyme in the Biosynthesis of L-Lysine By Plants And 'Chlamydia'

    SciTech Connect

    Watanabe, N.; Cherney, M.M.; van Belkum, M.J.; Marcus, S.L.; Flegel, M.D.; Clay, M.D.; Deyholos, M.K.; Vederas, J.C.; James, M.N.G.

    2007-07-13

    The essential biosynthetic pathway to l-Lysine in bacteria and plants is an attractive target for the development of new antibiotics or herbicides because it is absent in humans, who must acquire this amino acid in their diet. Plants use a shortcut of a bacterial pathway to l-Lysine in which the pyridoxal-5-phosphate (PLP)-dependent enzyme ll-diaminopimelate aminotransferase (LL-DAP-AT) transforms l-tetrahydrodipicolinic acid (L-THDP) directly to LL-DAP. In addition, LL-DAP-AT was recently found in Chlamydia sp., suggesting that inhibitors of this enzyme may also be effective against such organisms. In order to understand the mechanism of this enzyme and to assist in the design of inhibitors, the three-dimensional crystal structure of LL-DAP-AT was determined at 1.95 Angstroms resolution. The cDNA sequence of LL-DAP-AT from Arabidopsis thaliana (AtDAP-AT) was optimized for expression in bacteria and cloned in Escherichia coli without its leader sequence but with a C-terminal hexahistidine affinity tag to aid protein purification. The structure of AtDAP-AT was determined using the multiple-wavelength anomalous dispersion (MAD) method with a seleno-methionine derivative. AtDAP-AT is active as a homodimer with each subunit having PLP in the active site. It belongs to the family of type I fold PLP-dependent enzymes. Comparison of the active site residues of AtDAP-AT and aspartate aminotransferases revealed that the PLP binding residues in AtDAP-AT are well conserved in both enzymes. However, Glu97* and Asn309* in the active site of AtDAP-AT are not found at similar positions in aspartate aminotransferases, suggesting that specific substrate recognition may require these residues from the other monomer. A malate-bound structure of AtDAP-AT allowed LL-DAP and L-glutamate to be modeled into the active site. These initial three-dimensional structures of LL-DAP-AT provide insight into its substrate specificity and catalytic mechanism.

  19. A Method to Determine Lysine Acetylation Stoichiometries

    DOE PAGES

    Nakayasu, Ernesto S.; Wu, Si; Sydor, Michael A.; ...

    2014-01-01

    Lysine acetylation is a common protein posttranslational modification that regulates a variety of biological processes. A major bottleneck to fully understanding the functional aspects of lysine acetylation is the difficulty in measuring the proportion of lysine residues that are acetylated. Here we describe a mass spectrometry method using a combination of isotope labeling and detection of a diagnostic fragment ion to determine the stoichiometry of protein lysine acetylation. Using this technique, we determined the modification occupancy for ~750 acetylated peptides from mammalian cell lysates. Furthermore, the acetylation on N-terminal tail of histone H4 was cross-validated by treating cells with sodiummore » butyrate, a potent deacetylase inhibitor, and comparing changes in stoichiometry levels measured by our method with immunoblotting measurements. Of note we observe that acetylation stoichiometry is high in nuclear proteins, but very low in mitochondrial and cytosolic proteins. In summary, our method opens new opportunities to study in detail the relationship of lysine acetylation levels of proteins with their biological functions.« less

  20. The effect of condensed tannins in Lotus corniculatus on plasma metabolism of methionine, cystine and inorganic sulphate by sheep.

    PubMed

    Wang, Y; Waghorn, G C; Barry, T N; Shelton, I D

    1994-12-01

    Fresh Lotus corniculatus containing 27 g extractable condensed tannin (CT)/kg dry matter (DM) and 8 g bound CT/kg DM was fed at hourly intervals to sheep held in metabolism cages to study the effects of CT on nutrient digestion and on metabolism of methionine, cystine and inorganic sulphate in plasma. Polyethylene glycol (PEG) was continuously infused into the rumen of half the sheep to remove the effects of CT. Principal measurements in the two groups were plasma irreversible loss (IRL) rate and interconversions of methionine, cystine and inorganic sulphate using 35S labelling. CT in Lotus corniculatus had no effects on the apparent digestion of cellulose and minerals, slightly depressed DM, organic matter and hemicellulose digestion and markedly reduced the apparent digestion of N (P < 0.01). The concentration of NH3 and molar proportions of iso-butyric acid, iso-valeric acid and n-valeric acid in rumen fluid were markedly increased by the PEG infusion (P < 0.01), whereas total volatile fatty acid concentration and molar proportions of acetic acid, propionic acid and n-butyric acid were not affected. PEG infusion temporarily increased rumen protozoa numbers. CT greatly increased the IRL of plasma cystine (13.1 v. 7.0 mumol/min; P < 0.05) and reduced IRL of plasma inorganic sulphate (36.8 v. 48.1 mumol/min; P < 0.01) but had no effect on methionine IRL. CT increased transulphuration of methionine to cystine (4.37 v. 1.24 mumol/min; P < 0.05), increased cystine entering the plasma from whole-body protein turnover plus absorption from the small intestine (9.34 v. 5.75 mumol/min; P < 0.05) and increased cystine flux to body synthetic reactions (11.89 v. 5.41 mumol/min; P < 0.05). CT had no effect on the proportion of methionine total flux transferred to sulphate (0.05 v. 0.06; P < 0.05), reduced the proportion of methionine flux transferred to body synthetic reactions (0.68 v. 0.86) and markedly reduced the proportion of cystine flux transferred to sulphate (0.09 v

  1. Alimentary proteins, amino acids and cholesterolemia.

    PubMed

    Blachier, François; Lancha, Antonio H; Boutry, Claire; Tomé, Daniel

    2010-01-01

    Numerous data from both epidemiological and experimental origins indicate that some alimentary proteins and amino acids in supplements can modify the blood LDL cholesterol, HDL cholesterol and total cholesterol. After an initial approval of the health claim for soy protein consumption for the prevention of coronary heart disease, more recently it has been concluded from an overall analysis of literature that isolated soy protein with isoflavones only slightly decrease LDL and total cholesterol. Other plant extracts and also some proteins from animal origin have been reported to exert a lowering effect on blood cholesterol when compared with a reference protein (often casein). The underlying mechanisms are still little understood. Individual amino acids and mixture of amino acids have also been tested (mostly in animal studies) for their effects on cholesterol parameters and on cholesterol metabolism. Methionine, lysine, cystine, leucine, aspartate and glutamate have been tested individually and in combination in different models of either normo or hypercholesterolemic animals and found to be able to modify blood cholesterol and/or LDL cholesterol and/or HDL cholesterol. It is however not known if these results are relevant to human nutrition.

  2. Oxidation of apolipoprotein(a) inhibits kringle-associated lysine binding: the loss of intrinsic protein fluorescence suggests a role for tryptophan residues in the lysine binding site.

    PubMed Central

    Hermann, A.; Laws, W. R.; Harpel, P. C.

    1997-01-01

    Lipoprotein(a) [Lp(a)] is a low-density lipoprotein complex consisting of apolipoprotein(a) [apo(a)] disulfide-linked to apolipoprotein B-100. Lp(a) has been implicated in atherogenesis and thrombosis through the lysine binding site (LBS) affinity of its kringle domains. We have examined the oxidative effect of 2,2'-azobis-(amidinopropane) HCl (AAPH), a mild hydrophilic free radical initiator, upon the ability of Lp(a) and recombinant apo(a), r-apo(a), to bind through their LBS domains. AAPH treatment caused a time-dependent decrease in the number of functional Lp(a) or r-apo(a) molecules capable of binding to fibrin or lysine-Sepharose and in the intrinsic protein fluorescence of both Lp(a) and r-apo(a). The presence of a lysine analogue during the reaction prevented the loss of lysine binding and provided a partial protection from the loss of tryptophan fluorescence. The partial protection of fluorescence by lysine analogues was observed in other kringle-containing proteins, but not in proteins lacking kringles. No significant aggregation, fragmentation, or change in conformation of Lp(a) or r-apo(a) was observed as assessed by native or SDS-PAGE, light scattering, retention of antigenicity, and protein fluorescence emission spectra. Our results suggest that AAPH destroys amino acids in the kringles of apo(a) that are essential for lysine binding, including one or more tryptophan residues. The present study, therefore, raises the possibility that the biological roles of Lp(a) may be mediated by its state of oxidation, especially in light of our previous study showing that the reductive properties of sulfhydryl-containing compounds increase the LBS affinity of Lp(a) for fibrin. PMID:9385634

  3. Genetic Studies of Sulfadiazine-resistant and Methionine-requiring Neisseria Isolated From Clinical Material

    PubMed Central

    Catlin, B. Wesley

    1967-01-01

    Deoxyribonucleate (DNA) preparations were extracted from Neisseria meningitidis (four isolates from spinal fluid and blood) and N. gonorrhoeae strains, all of which were resistant to sulfadiazine upon primary isolation. These DNA preparations, together with others from in vitro mutants of N. meningitidis and N. perflava, were examined in transformation tests by using as recipient a drug-susceptible strain of N. meningitidis (Ne 15 Sul-s Met+) which was able to grow in a methionine-free defined medium. The sulfadiazine resistance typical of each donor was introduced into the uniform constitution of this recipient. Production of p-aminobenzoic acid was not significantly altered thereby. Transformants elicited by DNA from the N. meningitidis clinical isolates were resistant to at least 200 μg of sulfadiazine/ml, and did not show a requirement for methionine (Sul-r Met+). DNA from six strains of N. gonorrhoeae, which were isolated during the period of therapeutic use of sulfonamides, conveyed lower degrees of resistance and, invariably, a concurrent methionine requirement (Sul-r/Met−). The requirement of these transformants, and that of in vitro mutants selected on sulfadiazine-agar, was satisfied by methionine, but not by vitamin B12, homocysteine, cystathionine, homoserine, or cysteine. Sul-r Met+ and Sul-r/Met− loci could coexist in the same genome, but were segregated during transformation. On the other hand, the dual Sul-r/Met− properties were not separated by recombination, but were eliminated together. DNA from various Sul-r/Met− clones tested against recipients having nonidentical Sul-r/Met− mutant sites yielded Sul-s Met+ transformants. The met locus involved is genetically complex, and will be a valuable tool for studies of genetic fine structure of members of Neisseria, and of genetic homology between species. Images PMID:4962305

  4. Fragmentation network of doubly charged methionine: Interpretation using graph theory.

    PubMed

    Ha, D T; Yamazaki, K; Wang, Y; Alcamí, M; Maeda, S; Kono, H; Martín, F; Kukk, E

    2016-09-07

    The fragmentation of doubly charged gas-phase methionine (HO2CCH(NH2)CH2CH2SCH3) is systematically studied using the self-consistent charge density functional tight-binding molecular dynamics (MD) simulation method. We applied graph theory to analyze the large number of the calculated MD trajectories, which appears to be a highly effective and convenient means of extracting versatile information from the large data. The present theoretical results strongly concur with the earlier studied experimental ones. Essentially, the dication dissociates into acidic group CO2H and basic group C4NSH10. The former may carry a single or no charge and stays intact in most cases, whereas the latter may hold either a single or a double charge and tends to dissociate into smaller fragments. The decay of the basic group is observed to follow the Arrhenius law. The dissociation pathways to CO2H and C4NSH10 and subsequent fragmentations are also supported by ab initio calculations.

  5. 21 CFR 172.399 - Zinc methionine sulfate.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... ADDITIVES PERMITTED FOR DIRECT ADDITION TO FOOD FOR HUMAN CONSUMPTION Special Dietary and Nutritional... reaction between equimolar amounts of zinc sulfate and DL-methionine in purified water. (b) The...

  6. Appropriate statistical methods to compare dose responses of methionine sources.

    PubMed

    Kratzer, D D; Littell, R C

    2006-05-01

    Two sources of methionine (Met) activity are frequently used in commercial feed formulation: DL-2-hydroxy-4-(methylthio) butanoic acid (HMTBA), most commonly available as an 88% solution with 12% water; and DL-methionine (DLM, 99% powder). Despite the fact that both compounds have been in commercial use for over 50 yr, controversy and confusion remain with respect to their relative bioefficacy (RBE). This paper presents a review of the use of a nonlinear common plateau asymptotic regression technique (NLCPAR) that has been used to compare the 2 Met sources with particular emphasis on the validity of the basic assumptions of that model. The thesis of this paper is that the controversy is due, at least in part, to the misapplication of this regression technique to estimate the RBE of HMTBA and DLM. The NLCPAR model is a bioassay with the key dependent assumptions that HMTBA is a dilution of DLM, and that each follows dose-response curves of the same form and approach a common plateau. Because both provide Met activity, it may be considered reasonable to accept these assumptions; however, specifically testing them demonstrated that the assumption of a common dose-response is not supported by data. The common plateau assumption was tested with an alternative approach of fitting nonlinear separate plateaus asymptotic regression (NLSPAR) to a set of 13 published broiler studies in which the NLCPAR model had been used to estimate RBE of HMTBA and DLM. The hypothesis of a common plateau was rejected (P < 0.01), meaning that the conclusion that HMTBA had lower bioefficacy than DLM based on the NLCPAR methodology was not valid. An example using published data demonstrated that the NLSPAR model was a significantly better fit than the NLCPAR model, and showed that HMTBA and DLM followed different dose responses. Consequently, there was no single value for RBE for the entire dose range; rather, the RBE of the 2 compounds varied with use level. The evidence presented here

  7. Dendronized nanoconjugates of lysine and folate for treatment of cancer.

    PubMed

    Jain, Keerti; Gupta, Umesh; Jain, Narendra K

    2014-08-01

    Poly-L-lysine (PLL) dendrimers are currently being investigated as antiangiogenic agent for therapy of cancer. In this study, we report folate conjugated poly-l-lysine dendrimers (FPLL) as an efficient carrier for model anticancer drug, doxorubicin hydrochloride (Dox); for pH sensitive drug release, selective targeting to cancer cells, anticancer activity and antiangiogenic activity. This nanoconjugate of Dox showed initial rapid in vitro release followed by gradual slow release, and the drug release was found to be pH sensitive with greater release at acidic pH. In the CAM assay and tubule formation assay with HUVEC, Dox-FPLL formulation showed the significant antiangiogenic activity confirming that activity of PLL was not compromised by the presence of Dox and folic acid. The ex vivo investigations with human breast cancer cell lines MCF-7 showed enhanced cytotoxicity of Dox-FPLL with significantly enhanced intracellular uptake (p<0.001). The in vivo therapeutic potential of nanoconjugate was determined in MCF-7 breast cancer xenograft model in tumor-bearing mice. Dox-FPLL increased the concentration of Dox in tumor by 121.5-fold after 24 h in comparison with free Dox formulation. The folate conjugated dendrimeric Dox showed superior anti-tumor activity in tumor xenograft model with significantly prolonged survival determined by Kaplan Meier survival analysis (p<0.001).

  8. Pre-steady-state kinetic and structural analysis of interaction of methionine γ-lyase from Citrobacter freundii with inhibitors.

    PubMed

    Kuznetsov, Nikita A; Faleev, Nicolai G; Kuznetsova, Alexandra A; Morozova, Elena A; Revtovich, Svetlana V; Anufrieva, Natalya V; Nikulin, Alexei D; Fedorova, Olga S; Demidkina, Tatyana V

    2015-01-02

    Methionine γ-lyase (MGL) catalyzes the γ-elimination of l-methionine and its derivatives as well as the β-elimination of l-cysteine and its analogs. These reactions yield α-keto acids and thiols. The mechanism of chemical conversion of amino acids includes numerous reaction intermediates. The detailed analysis of MGL interaction with glycine, l-alanine, l-norvaline, and l-cycloserine was performed by pre-steady-state stopped-flow kinetics. The structure of side chains of the amino acids is important both for their binding with enzyme and for the stability of the external aldimine and ketimine intermediates. X-ray structure of the MGL·l-cycloserine complex has been solved at 1.6 Å resolution. The structure models the ketimine intermediate of physiological reaction. The results elucidate the mechanisms of the intermediate interconversion at the stages of external aldimine and ketimine formation.

  9. Growth rate of Enterobacteriaceae at elevated temperatures: limitation by methionine.

    PubMed

    Ron, E Z

    1975-10-01

    The effect of elevated temperatures on growth rate was studied in five strains of Enterobacteriaceae. In all the strains tested a shift to the elevated temperature resulted in an immediate decrease in growth rate which was due to limitation in the availability of endogenous methionine. The first biosynthetic enzyme of the methionine pathway-homoserine transsuccinylase-was studied in extracts of Aerobacter aerogenes, Salmonella typhimurium, and Escherichia coli and was shown to be temperature sensitive in all of them.

  10. Loss of conformational stability in calmodulin upon methionine oxidation.

    PubMed Central

    Gao, J; Yin, D H; Yao, Y; Sun, H; Qin, Z; Schöneich, C; Williams, T D; Squier, T C

    1998-01-01

    We have used electrospray ionization mass spectrometry (ESI-MS), circular dichroism (CD), and fluorescence spectroscopy to investigate the secondary and tertiary structural consequences that result from oxidative modification of methionine residues in wheat germ calmodulin (CaM), and prevent activation of the plasma membrane Ca-ATPase. Using ESI-MS, we have measured rates of modification and molecular mass distributions of oxidatively modified CaM species (CaMox) resulting from exposure to H2O2. From these rates, we find that oxidative modification of methionine to the corresponding methionine sulfoxide does not predispose CaM to further oxidative modification. These results indicate that methionine oxidation results in no large-scale alterations in the tertiary structure of CaMox, because the rates of oxidative modification of individual methionines are directly related to their solvent exposure. Likewise, CD measurements indicate that methionine oxidation results in little change in the apparent alpha-helical content at 28 degrees C, and only a small (0.3 +/- 0.1 kcal mol(-1)) decrease in thermal stability, suggesting the disruption of a limited number of specific noncovalent interactions. Fluorescence lifetime, anisotropy, and quenching measurements of N-(1-pyrenyl)-maleimide (PMal) covalently bound to Cys26 indicate local structural changes around PMal in the amino-terminal domain in response to oxidative modification of methionine residues in the carboxyl-terminal domain. Because the opposing globular domains remain spatially distant in both native and oxidatively modified CaM, the oxidative modification of methionines in the carboxyl-terminal domain are suggested to modify the conformation of the amino-terminal domain through alterations in the structural features involving the interdomain central helix. The structural basis for the linkage between oxidative modification and these global conformational changes is discussed in terms of possible alterations in

  11. Enhancement of ε-poly-L-lysine production coupled with precursor L-lysine feeding in glucose-glycerol co-fermentation by Streptomyces sp. M-Z18.

    PubMed

    Chen, Xu-Sheng; Ren, Xi-Dong; Zeng, Xin; Zhao, Fu-Lin; Tang, Lei; Zhang, Hong-Jian; Zhang, Jian-Hua; Mao, Zhong-Gui

    2013-12-01

    ε-Poly-L-lysine (ε-PL), one of the only two homo-poly amino acids known in nature, is used as a preservative. In this study, strategies of feeding precursor L-lysine into 5 L laboratory scale fermenters, including optimization of L-lysine concentration and time, was investigated to optimize the production of ε-PL by Streptomyces sp. M-Z18. The optimized strategy was then used in ε-PL fed-batch fermentation in which glucose and glycerol served as mixed carbon sources. In this way, a novel ε-PL production strategy involving precursor L-lysine coupled with glucose-glycerol co-fermentation was developed. Under optimal conditions, ε-PL production reached 37.6 g/l, which was 6.2 % greater than in a previous study in which glucose and glycerol co-fermentation was performed without added L-lysine (35.14 g/l). To the best of our knowledge, this is the first report of the enhancement of ε-PL production through L-lysine feeding to evaluate the use of fermenters. Meanwhile, the role of L-lysine in the promotion of ε-PL production, participating ε-PL synthesis as a whole, was first determined using the L-[U-(13)C] lysine labeling method. It has been suggested that the bottleneck of ε-PL synthesis in Streptomyces sp. M-Z18 is in the biosynthesis of precursor L-lysine. The information obtained in the present work may facilitate strain improvement and efficient large-scale ε-PL production.

  12. Amino acid composition of Lagenaria siceraria seed flour and protein fractions.

    PubMed

    Ogunbusola, Moriyike Esther; Fagbemi, Tayo Nathaniel; Osundahunsi, Oluwatooyin Faramade

    2010-12-01

    Defatted seed flours of Lagenaria siceraria (calabash and bottle gourd) were fractionated into their major protein fractions. The amino acid composition of seed flours and their protein fractions were determined and the protein quality was evaluated. Glutamic acid (139-168 mg/g protein) was the most abundant amino acid followed by aspartic acid (89.0-116 mg/g protein) in both the seed flours and their protein fractions. The total essential amino acid ranged from 45.8 to 51.5%. The predicted protein efficiency ratio and the predicted biological value ranged from 2.4 to 2.9 and 8.7 to 44.0, respectively. Lysine and sulphur amino acids were mostly concentrated in the globulin fractions. The first and second limiting amino acids in seed flours and protein fractions were methionine and valine or threonine. The seed flours contained adequate essential amino acids required by growing school children and adults. The seed has potential as protein supplement in cereal based complementary diets or in the replacement of animal proteins in conventional foods.

  13. Quantitative study in vivo of methionine cycle in humans using (methyl-/sup 2/H/sub 3/)- and (1-/sup 13/C)methionine

    SciTech Connect

    Storch, K.J.; Wagner, D.A.; Burke, J.F.; Young, V.R.

    1988-09-01

    Kinetic aspects of body methionine (MET) metabolism were examined in healthy young men during the fed and postabsorptive (PA) states. Rates of MET incorporation (S) into and release (B) from body proteins; transmethylation (TM); and remethylation (RM) and transsulfuration (TS) of homocysteine (HCY) were estimated with the aid of a 5-h constant intravenous infusion of (methyl-2H3)- and (1-13C)methionine. The isotopic data (plasma methionine labeling and 13C enrichment of expired air) were submitted to a stochastic model of amino acid metabolism. During the fed state, the subjects (n = 4) received, at 20-min intervals, small isonitrogenous isocaloric meals containing a complete L-amino acid mixture supplying MET at a rate equivalent to 198 mumol.kg body wt-1.day-1. The PA subjects (n = 4) received the isotope after a 10-h overnight fast. For the PA group, the components of MET metabolism were as follows: S, 20 +/- 0.5; B, 24 +/- 0.5; TM, 5.8 +/- 0.6; RM, 1.8 +/- 0.4; and TS, 4.0 +/- 0.4 (+/-SE) mumol.kg-1.h-1. During the fed state the values were S, 26 +/- 2.5; B, 18 +/- 2; TM, 14 +/- 1.3; RM, 5.7 +/- 0.9; and TS 8.3 +/- 0.6 mumol.kg-1.h-1. The meal-induced changes in B, TM, RM, and TS were significant (P less than 0.05). Comparison of the partitioning of MET between S and TM (these two pathways of MET disposal constitute the ''methionine locus'') in the PA and in the fed states indicates that the MET locus is of regulatory importance in MET homeostasis. A twofold increase in the partitioning of MET to TM was observed in the fed state. The increase in HCY recycling, relative to TS (these two pathways of HCY disposal constitute the ''HCY locus''), in the fed state did not reach statistical significance when compared with the PA state. Total daily TM are estimated to be 238 +/- 22 mumol/kg. This is similar to the estimate generated by the methyl balance model of Mudd and Poole which approximated 241 mumol/kg.

  14. [NMR screening of potential inhibitors of Citrobacter freundii methionine].

    PubMed

    Batuev, E A; Lizunov, A Iu; Morozova, E A; Klochkov, V V; Anufrieva, N V; Demidkina, T V; Pol'shakov, V I

    2014-01-01

    Methionine γ-lyase [EC 4.4.1.11] participates in a methionine catabolism at a number of bacteria and protozoa eukaryotes, including pathogenic microorganisms. Lack of this enzyme at mammals allows consider it as a perspective target for rational antibacterial drug design. Currently in medical practice there are no the preparations based on an inhibition of methionine γ-lyase activity. We present results of the search of potential inhibitors of the enzyme using the NMR screening techniques based on identification of compounds, which able to bind specifically to their biological target. Study included a stage of in silico virtual screening of the library of commercially available compounds and subsequent experimental selection of the leading compounds, capable to interact with enzyme. Identification of binding was carried out by means of saturation transfer difference (STD) spectroscopy and WaterLOGSY technique. At the final stage the experimental assessment of inhibiting ability of the selected compounds in the reaction of γ-elimination of L-methionine catalyzed by methionine γ-lyase was carried out. Binding constants of two leading compounds were determined using the WaterLOGSY method. The research expands structural group of potential inhibitors of methionine γ-lyase and allows approach to the design of the inhibitors with higher efficacy.

  15. Modeling the oxidation of methionine residues by peroxides in proteins.

    PubMed

    Chennamsetty, Naresh; Quan, Yong; Nashine, Vishal; Sadineni, Vikram; Lyngberg, Olav; Krystek, Stanley

    2015-04-01

    We report the use of molecular modeling to predict the oxidation propensity of methionine residues in proteins. Oxidation of methionine to the sulfoxide form is one of the major degradation pathways for therapeutic proteins. Oxidation can occur during production, formulation, or storage of pharmaceuticals and it often reduces or eliminates biological activity. We use a molecular model based on atomistic simulations called 2-shell water coordination number to predict the oxidation rates for several model proteins and therapeutic candidates. In addition, we implement models that are based on static and simulation average of the solvent-accessible area (SAA) for either the side chain or the sulfur atom in the methionine residue. We then compare the results from the different models against the experimentally measured relative rates of methionine oxidation. We find that both the 2-shell model and the simulation-averaged SAA models are accurate in predicting the oxidation propensity of methionine residues for the proteins tested. We also find the appropriate parameter ranges where the models are most accurate. These models have significant predictive power and can be used to enable further protein engineering or to guide formulation approaches in stabilizing the unstable methionine residues.

  16. Elongation factor 1 alpha concentration is highly correlated with the lysine content of maize endosperm.

    PubMed Central

    Habben, J E; Moro, G L; Hunter, B G; Hamaker, B R; Larkins, B A

    1995-01-01

    Lysine is the most limiting essential amino acid in cereals, and for many years plant breeders have attempted to increase its concentration to improve the nutritional quality of these grains. The opaque2 mutation in maize doubles the lysine content in the endosperm, but the mechanism by which this occurs is unknown. We show that elongation factor 1 alpha (EF-1 alpha) is overexpressed in opaque2 endosperm compared with its normal counterpart and that there is a highly significant correlation between EF-1 alpha concentration and the total lysine content of the endosperm. This relationship is also true for two other cereals, sorghum and barley. It appears that genetic selection for genotypes with a high concentration of EF-1 alpha can significantly improve the nutritional quality of maize and other cereals. Images Fig. 1 Fig. 2 PMID:7567989

  17. A System for Enzymatic Lysine Methylation in a Desired Sequence Context

    PubMed Central

    2017-01-01

    A number of lysine-specific methyltransferases (KMTs) are responsible for the post-translational modification of cellular proteins on lysine residues. Most KMTs typically recognize specific motifs in unstructured, short peptide sequences. However, we have recently discovered a novel KMT that appeared to have a more relaxed sequence specificity, namely, valosin-containing protein (VCP)-KMT, which trimethylates Lys-315 in the molecular chaperone VCP. On the basis of this, here, we explored the possibility of using the VCP-KMT/VCP system to obtain specific lysine methylation of desired sequences grafted onto a VCP-derived scaffold. We generated VCP-derived proteins in which three amino acid residues on each side of Lys-315 had been replaced by various sequences representing lysine methylation sites in histone H3. We found that all of these chimeric proteins were subject to efficient VCP-KMT-mediated methylation in vitro, and methylation was also observed in mammalian cells. Thus, we here describe a versatile system for introducing lysine methylation into a desired peptide sequence, and the approach should be readily expandable for generating combinatorial libraries of methylated sequences. PMID:28357416

  18. Crystal structure of Mycobacterium tuberculosis diaminopimelate decarboxylase, an essential enzyme in bacterial lysine biosynthesis.

    PubMed

    Gokulan, Kuppan; Rupp, Bernhard; Pavelka, Martin S; Jacobs, William R; Sacchettini, James C

    2003-05-16

    The Mycobacterium tuberculosis lysA gene encodes the enzyme meso-diaminopimelate decarboxylase (DAPDC), a pyridoxal-5'-phosphate (PLP)-dependent enzyme. The enzyme catalyzes the final step in the lysine biosynthetic pathway converting meso-diaminopimelic acid (DAP) to l-lysine. The lysA gene of M. tuberculosis H37Rv has been established as essential for bacterial survival in immunocompromised mice, demonstrating that de novo biosynthesis of lysine is essential for in vivo viability. Drugs targeted against DAPDC could be efficient anti-tuberculosis drugs, and the three-dimensional structure of DAPDC from M. tuberculosis complexed with reaction product lysine and the ternary complex with PLP and lysine in the active site has been determined. The first structure of a DAPDC confirms its classification as a fold type III PLP-dependent enzyme. The structure shows a stable 2-fold dimer in head-to-tail arrangement of a triose-phosphate isomerase (TIM) barrel-like alpha/beta domain and a C-terminal beta sheet domain, similar to the ornithine decarboxylase (ODC) fold family. PLP is covalently bound via an internal aldimine, and residues from both domains and both subunits contribute to the binding pocket. Comparison of the structure with eukaryotic ODCs, in particular with a di-fluoromethyl ornithine (DMFO)-bound ODC from Trypanosoma bruceii, indicates that corresponding DAP-analogues might be potential inhibitors for mycobacterial DAPDCs.

  19. The lysine biosynthetic enzyme Lys4 influences iron metabolism, mitochondrial function and virulence in Cryptococcus neoformans.

    PubMed

    Do, Eunsoo; Park, Minji; Hu, Guanggan; Caza, Mélissa; Kronstad, James W; Jung, Won Hee

    2016-09-02

    The lysine biosynthesis pathway via α-aminoadipate in fungi is considered an attractive target for antifungal drugs due to its absence in mammalian hosts. The iron-sulfur cluster-containing enzyme homoaconitase converts homocitrate to homoisocitrate in the lysine biosynthetic pathway, and is encoded by LYS4 in the model yeast Saccharomyces cerevisiae. In this study, we identified the ortholog of LYS4 in the human fungal pathogen, Cryptococcus neoformans, and found that LYS4 expression is regulated by iron levels and by the iron-related transcription factors Hap3 and HapX. Deletion of the LYS4 gene resulted in lysine auxotrophy suggesting that Lys4 is essential for lysine biosynthesis. Our study also revealed that lysine uptake was mediated by two amino acid permeases, Aap2 and Aap3, and influenced by nitrogen catabolite repression (NCR). Furthermore, the lys4 mutant showed increased sensitivity to oxidative stress, agents that challenge cell wall/membrane integrity, and azole antifungal drugs. We showed that these phenotypes were due in part to impaired mitochondrial function as a result of LYS4 deletion, which we propose disrupts iron homeostasis in the organelle. The combination of defects are consistent with our observation that the lys4 mutant was attenuated virulence in a mouse inhalation model of cryptococcosis.

  20. Effect of irradiation on Nε-carboxymethyl-lysine and Nε-carboxyethyl-lysine formation in cooked meat products during storage

    NASA Astrophysics Data System (ADS)

    Yu, Ligang; He, Zhiyong; Zeng, Maomao; Zheng, Zongping; Chen, Jie

    2016-03-01

    This study investigated the effects of irradiation on Nε-carboxymethyl-lysine (CML) and Nε-carboxyethyl-lysine (CEL) formation in cooked red and white meats during storage. The results showed that irradiation did not affect CML/CEL formation (0 weeks). After 6 weeks, CML/CEL contents in the irradiated samples exhibited a higher growth rate than the non-irradiated samples, especially the red meat. The results of electron spin resonance spectrometry and 2-Thiobarbituric acid-reactive substances suggested irradiation had induced free-radical reactions and accelerated lipid oxidation during storage. A linear correlation (r=0.810-0.906, p<0.01) was found between the loss of polyunsaturated fatty acids content and increase of CML/CEL content in the irradiated samples after 0 and 6 weeks of storage. The results indicate that irradiation-induced lipid oxidation promotes CML/CEL formation, and CML/CEL formation by the lipid oxidation pathways may be an important pathway for CML/CEL accumulation in irradiated meat products during storage.

  1. Meta-analysis of lactation performance in dairy cows receiving supplemental dietary methionine sources or postruminal infusion of methionine.

    PubMed

    Zanton, G I; Bowman, G R; Vázquez-Añón, M; Rode, L M

    2014-11-01

    The objectives of our study were to evaluate the productive response to methionine supplementation in lactating dairy cows and to define a relationship between metabolizable Met (MP Met) intake and production. A database of 64 papers meeting the selection criteria was developed evaluating postruminally infused dl-methionine (9 papers with 18 control diets and 35 treatment comparisons), 2-hydroxy-4-methylthio butanoic acid (HMTBa) provided as either a liquid or Ca salt form (17 papers with 34 control diets and 46 treatment comparisons), Mepron (Evonik Industries, Essen, Germany; 18 papers with 35 control diets and 42 treatment comparisons), and Smartamine (Adisseo Inc., Antony, France; 20 papers with 30 control diets and 39 treatment comparisons). Dietary ingredients and their accompanying nutritional compositions as described in the reports were entered into the Cornell-Penn-Miner software to model the diets and to predict nutrients that were not reported in the original publication. Data were analyzed using a weighted analysis of response to supplementation compared with the intraexperiment control, as well as through a regression analysis to changing dietary MP Met. Data included in the analysis were from experiments published between 1970 and 2011 with cows supplemented with between 3.5 and 67.9 g of Met or its equivalent from HMTBa. Cows supplemented with Smartamine consumed more, whereas cows supplemented with Mepron consumed less DM compared with controls. Milk yield did not significantly respond to Met supplementation, although it tended to increase for cows supplemented with HMTBa and Mepron. Milk protein yield was increased due to supplementation from all sources or from infusion, and protein concentration was greater for all supplements or infusion of dl-Met, except for cows supplemented with HMTBa. Irrespective of Met source, milk protein yield increased 2.23 g of protein/g of MP Met until reaching the breakpoint. Milk fat yield was increased for Mepron

  2. Abundance and functional roles of intrinsic disorder in the antimicrobial peptides of the NK-lysin family.

    PubMed

    Yacoub, Haitham A; Al-Maghrabi, Omar A; Ahmed, Ekram S; Uversky, Vladimir N

    2017-03-01

    NK-lysins are antimicrobial peptides (AMPs) that participate in the innate immune response and also have several pivotal roles in various biological processes. Such multifunctionality is commonly found among intrinsically disordered proteins. However, NK-lysins have never been systematically analyzed for intrinsic disorder. To fill this gap, the amino acid sequences of NK-lysins from various species were collected from UniProt and used for the comprehensive computational analysis to evaluate the propensity of these proteins for intrinsic disorder and to investigate the potential roles of disordered regions in NK-lysin functions. We analyzed abundance and peculiarities of intrinsic disorder distribution in all-known NK-lysins and showed that many NK-lysins are expected to have substantial levels of intrinsic disorder. Curiously, high level of intrinsic disorder was also found even in two proteins with known 3D-strucutres (NK-lysin from pig and human granulysin). Many of the identified disordered regions can be involved in protein-protein interactions. In fact, NK-lysins are shown to contain three to eight molecular recognition features; i.e. short structure-prone segments which are located within the long disordered regions and have a potential to undergo a disorder-to-order transition upon binding to a partner. Furthermore, these disordered regions are expected to have several sites of various posttranslational modifications. Our study shows that NK-lysins, which are AMPs with a set of prominent roles in the innate immune response, are expected to abundantly possess intrinsically disordered regions that might be related to multifunctionality of these proteins in the signal transduction pathways controlling the host response to pathogenic agents.

  3. N-formylation of lysine in histone proteins as a secondary modification arising from oxidative DNA damage.

    PubMed

    Jiang, Tao; Zhou, Xinfeng; Taghizadeh, Koli; Dong, Min; Dedon, Peter C

    2007-01-02

    The posttranslational modification of histone and other chromatin proteins has a well recognized but poorly defined role in the physiology of gene expression. With implications for interfering with these epigenetic mechanisms, we now report the existence of a relatively abundant secondary modification of chromatin proteins, the N(6)-formylation of lysine that appears to be uniquely associated with histone and other nuclear proteins. Using both radiolabeling and sensitive bioanalytical methods, we demonstrate that the formyl moiety of 3'-formylphosphate residues arising from 5'-oxidation of deoxyribose in DNA, caused by the enediyne neocarzinostatin, for example, acylate the N(6)-amino groups of lysine side chains. A liquid chromatography (LC)-tandem mass spectrometry (MS) method was developed to quantify the resulting N(6)-formyl-lysine residues, which were observed to be present in unperturbed cells and all sources of histone proteins to the extent of 0.04-0.1% of all lysines in acid-soluble chromatin proteins including histones. Cells treated with neocarzinostatin showed a clear dose-response relationship for the formation of N(6)-formyl-lysine, with this nucleosome linker-selective DNA-cleaving agent causing selective N(6)-formylation of the linker histone H1. The N(6)-formyl-lysine residue appears to represent an endogenous histone secondary modification, one that bears chemical similarity to lysine N(6)-acetylation recognized as an important determinant of gene expression in mammalian cells. The N(6)-formyl modification of lysine may interfere with the signaling functions of lysine acetylation and methylation and thus contribute to the pathophysiology of oxidative and nitrosative stress.

  4. Uptake of L-lysine by a double mutant of Saccharomyces cerevisiae.

    PubMed

    García, J C; Kotyk, A

    1988-01-01

    A gap1 can1 mutant of Saccharomyces cerevisiae with a single lysine transport system remaining was used to study detailed kinetics of this transport. Its half-saturation constant was 78 mumol per litre, its maximum rate of transport was 0.29 mumol L-lysine per g dry matter per minute, both parameters being lower by more than an order of magnitude in comparison with the GAP system. The pH optimum lay at very acid values of about 3, the temperature dependence without any transition point showed an activation energy of 48 kJ/mol. The transport was inhibited by common metabolic inhibitors (3'-chlorophenylhydrazonomalononitrile, antimycin, 2-deoxy-D-glucose, sodium arsenate) as well as by a membrane-active one (uranyl nitrate). The specificity of the system was extremely high, none of the natural amino acids acting as competitor to L-lysine. The maximum accumulation ratio attained (at about 5 mg dry matter per mL) was 100: 1-120: 1, in agreement with the measured protonmotive force under the assumption of 1 H+ ion being transported with 1 lysine molecule. The ratio decreased with increasing external concentration of lysine to as little as 4: 1 at 1 mmol lysine per litre. It also decreased with increasing suspension density and it was at extremely low suspension densities (0.2 mg dry matter per mL) that ratios of as much as 500: 1 were reached. Application of group-specific inhibitors showed that the active site of the carrier contains an essential histidine residue.

  5. N. sup. var epsilon. -acetyl-. beta. -lysine: An osmolyte synthesized by mothanogenic archaebacteria

    SciTech Connect

    Sowers, K.R.; Gunsalus, R.P. ); Robertson, D.E.; Noll, D.; Roberts, M.F. )

    1990-12-01

    Methanosarcina thermophila, a nonmarine methanogenic archaebacterium, can grow in a range of saline concentrations. At less than 0.4 M NaCl, Ms. thermophila accumulated glutamate in response to increasing osmotic stress. At greater than 0.4 M NaCl, this organism synthesized a modified {beta}-amino acid that was identified as N{sup {var epsilon}}-acetyl-{beta}-lysine by NMR spectroscopy and ion-exchange HPLC. This {beta}-amino acid derivative accumulated to high intracellular concentrations (up to 0.6 M) in Ms. thermophila and in another methanogen examined - Methanogenium cariaci, a marine species. The compound has features that are characteristic of a compatible solute: it is neutrally charged at physiological pH and it is highly soluble. When the cells were grown in the presence of exogenous glycine betaine, a physiological pH and it is highly soluble. When the cells were grown in the presence of exogenous glycine betaine, a physiological compatible solute, N{sup {var epsilon}}-acetyl-{beta}-lysine synthesis was repressed and glycine betaine was accumulated. N{sup {var epsilon}}-Acetyl-{beta}-lysine was synthesized by species from three phylogenetic families when grown in high solute concentrations, suggesting that it may be ubiquitous among the methanogens. The ability to control the biosynthesis of N{sup {var epsilon}}-acetyl-{beta}-lysine in response to extracellular solute concentration indicates that the methanogenic archaebacteria have a unique {beta}-amino acid biosynthetic pathway that is osmotically regulated.

  6. Lauryl-poly-L-lysine: A New Antimicrobial Agent?

    PubMed Central

    Thuault, Véronique; Mangas, Arturo; Thienpont, Anne; Geffard, Michel

    2014-01-01

    The development of multiple antibiotic resistance is a global problem. It is necessary to find new tools whose mechanisms of action differ from those of currently used antibiotics. It is known that fatty acids and cationic polypeptides are able to fight bacteria. Here, we describe the synthesis of fatty acids linked to a polypeptide with antibacterial activity. The linkage of fatty acids to a polypeptide is reported to increase the antibacterial effect of the linked fatty acid in comparison with free fatty acids (FA) or free poly-L-lysine (PLL) or a mixture of both (FA free + PLL free). A number of C6–C18 fatty acids were linked to PLL to obtain new synthetic products. These compounds were assessed in vitro to evaluate their antibacterial activity. Some fatty acid-PLLs showed a good ability to fight bacteria. Their bactericidal activity was evaluated, and, lauryl linked to PLL was found to be the most active product against both Gram-positive and Gram-negative bacteria. This new active component showed a good degree of specificity and reproducibility and its minimum inhibitory concentration (MIC) was comparatively good. The antibacterial activity of the lauryl-PLL compound suggests that it is a new and promising antimicrobial agent. PMID:24660058

  7. Toxicity of seleno-l-methionine, seleno-dl-methionine, high selenium wheat, and selenized yeast to mallard ducklings

    USGS Publications Warehouse

    Heinz, G.H.; Hoffman, D.J.; LeCaptain, L.J.

    1996-01-01

    The toxicity of four chemical forms of selenium (seleno-L-methionine, seleno-DL-methionine, selenized yeast, and high selenium wheat) was compared in day-old mallard ducklings (Anas platyrhynchos). In the first experiment, in which the basal diet was 75% wheat, survival after 2 weeks was lower for ducklings fed 30 ?g/g selenium as seleno-L-methionine (36%) than for ducklings fed 30 ?g/g selenium as seleno-DL-methionine (100%) or 30 ?g/g selenium from high selenium yeast (88%). In a second experiment, in which the basal diet was a commercial duck feed, survival after 2 weeks was 100% in ducklings fed 30 ?g/g selenium as seleno-DL-methionine, seleno-L-methionine, or selenized yeast. The greater toxicity of the L form of selenomethionine was probably related to the palatability or nutritional nature of the wheat-based diet used in experiment 1, but the exact reason for the difference between the DL and L forms is unknown. Biologically incorporated selenium, derived from high selenium wheat was no more toxic than selenium derived from the two purified forms of selenomethionine, and the selenium in selenized yeast was not as toxic as that in the two forms of selenomethionine.

  8. Analysis of Methionine Oxidation in Myosin Isoforms in Porcine Skeletal Muscle by LC-MS/MS Analysis

    PubMed Central

    Jeong, Jin-Yeon; Jung, Eun-Young; Jeong, Tae-Chul; Yang, Han-Sul; Kim, Gap-Don

    2016-01-01

    The purpose of this study was to analyze oxidized methionines in the myosin isoforms of porcine longissimus thoracis, psoas major, and semimembranosus muscles by liquid chromatography (LC) and mass spectrometry (MS). A total of 836 queries matched to four myosin isoforms (myosin-1, -2, -4, and -7) were analyzed and each myosin isoform was identified by its unique peptides (7.3-13.3). Forty-four peptides were observed from all three muscles. Seventeen peptides were unique to the myosin isoform and the others were common peptides expressed in two or more myosin isoforms. Five were identified as oxidized peptides with one or two methionine sulfoxides with 16 amu of mass modification. Methionines on residues 215 (215), 438 (438), 853 (851), 856 (854), 1071 (1069), and 1106 (1104) of myosin-1 (myosin-4) were oxidized by the addition of oxygen. Myosin-2 had two oxidized methionines on residues 215 and 438. No queries matched to myosin-7 were observed as oxidized peptides. LC-MS/MS allows analysis of the oxidation of specific amino acids on specific residue sites, as well as in specific proteins in the food system. PMID:27194935

  9. Effects of dietary sulfur amino acids on lead toxicity in chicks

    SciTech Connect

    Latta, D.M.

    1986-01-01

    Factorial experiments were conducted to examine the interactions of dietary methionine with cystine, choline and glycine in lead (Pb) intoxicated chicks. In Experiment 1, chicks were fed a basal diet deficient in methionine and total sulfur-containing amino acids (SAA) with 0 or 1000 ppm added Pb. Methionine or methionine plus cystine improved growth regardless of Pb level; cystine addition alone improved growth only when Pb was present. Dietary methionine appeared to counteract Pb toxicity more effectively than cystine. In Experiments 2 and 3 dietary variables were 0 or 1000 ppm Pb, adequate or inadequate methionine and marginal or excess (Experiment 2) or adequate or inadequate (Experiment 3) choline. In Experiment 2 growth depression by Pb was less with methionine-adequate compared to methionine-inadequate diets; there were no differences in growth with choline-marginal or choline-excess diets. In Experiment 3, the Pb-induced growth depression was exacerbated by adequate choline when methionine-inadequate diets were fed. It appears that Pb lowers the chick's choline requirement and that the methyl moiety of methionine does not participate directly in Pb detoxification. In Experiment 4 effects of adequate or deficient methionine, adequate or excess glycine and 0 or 1000 ppm Pb in choline-deficient chicks were studied. Methionine stimulated growth and the response was greater when excess glycine was present. Excess glycine stimulated growth only in the presence of adequate methionine suggesting glycine is limiting for growth in choline-deficient, methionine-adequate diets. These studies indicate that adequate methionine ameliorates Pb-induced growth depression in growing chicks but that the methionine effect is greater with choline-adequate than with choline-deficient diets. The amelioration of Pb toxicity by methionine may be partly related to increased excretion of Pb.

  10. 21 CFR 582.5411 - Lysine.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... 21 Food and Drugs 6 2012-04-01 2012-04-01 false Lysine. 582.5411 Section 582.5411 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) ANIMAL DRUGS, FEEDS, AND RELATED PRODUCTS SUBSTANCES GENERALLY RECOGNIZED AS SAFE Nutrients and/or Dietary Supplements...

  11. 21 CFR 582.5411 - Lysine.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... 21 Food and Drugs 6 2011-04-01 2011-04-01 false Lysine. 582.5411 Section 582.5411 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) ANIMAL DRUGS, FEEDS, AND RELATED PRODUCTS SUBSTANCES GENERALLY RECOGNIZED AS SAFE Nutrients and/or Dietary Supplements...

  12. 21 CFR 582.5411 - Lysine.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 21 Food and Drugs 6 2010-04-01 2010-04-01 false Lysine. 582.5411 Section 582.5411 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) ANIMAL DRUGS, FEEDS, AND RELATED PRODUCTS SUBSTANCES GENERALLY RECOGNIZED AS SAFE Nutrients and/or Dietary Supplements...

  13. 21 CFR 582.5411 - Lysine.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... 21 Food and Drugs 6 2013-04-01 2013-04-01 false Lysine. 582.5411 Section 582.5411 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) ANIMAL DRUGS, FEEDS, AND RELATED PRODUCTS SUBSTANCES GENERALLY RECOGNIZED AS SAFE Nutrients and/or Dietary Supplements...

  14. 21 CFR 582.5411 - Lysine.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... 21 Food and Drugs 6 2014-04-01 2014-04-01 false Lysine. 582.5411 Section 582.5411 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) ANIMAL DRUGS, FEEDS, AND RELATED PRODUCTS SUBSTANCES GENERALLY RECOGNIZED AS SAFE Nutrients and/or Dietary Supplements...

  15. Radioactive Lysine in Protein Metabolism Studies

    DOE R&D Accomplishments Database

    Miller, L. L.; Bale, W. F.; Yuile, C. L.; Masters, R. E.; Tishkoff, G. H.; Whipple,, G. H.

    1950-01-09

    Studies of incorporation of DL-lysine in various body proteins of the dog; the time course of labeled blood proteins; and apparent rate of disappearance of labeled plasma proteins for comparison of behavior of the plasma albumin and globulin fractions; shows more rapid turn over of globulin fraction.

  16. The Safety and Efficacy of Lysine Analogues in Cancer Patients: A Systematic Review and Meta-Analysis.

    PubMed

    Montroy, Joshua; Fergusson, Nicholas A; Hutton, Brian; Lavallée, Luke T; Morash, Chris; Cagiannos, Ilias; Cnossen, Sonya; Fergusson, Dean A; Breau, Rodney H

    2017-03-22

    Lysine analogues are effective agents used for the reduction of blood loss and transfusion. However, the safety of lysine analogues in cancer patients remains in question due to a potential risk of venous thromboembolism (VTE). The objective of our review is to investigate safety and efficacy of lysine analogue administration in the patients with cancer. Medline, Embase, and The Cochrane Library were searched from inception to June, 2016. Reference lists of retrieved studies were searched to identify additional publications. We included randomized clinical trials in adult cancer patients for which a lysine analogue was administered for the purpose of blood loss reduction. Abstract and full-text selection as well as data extraction and risk of bias assessment was done by 2 independent reviewers. The primary outcome was venous thromboembolic events. Secondary outcomes were other adverse events, blood transfusion, and blood loss. Overall, 11studies involving 1177 patients evaluated at least one of the primary or secondary outcomes. Nine studies evaluated the effects of tranexamic acid, one study evaluated the effects of aminocaproic acid and one study examined both agents. No increased risk of venous thromboembolism was observed for patients who received lysine analogues compared to control (Peto OR 0.58; 95% CI 0.26-1.28). The administration of a lysine analogue significantly decreased both transfusion risk (pooled RR 0.52, 95% CI 0.34-0.80) and blood loss (SMD -1.57, 95% CI -2.21 to -0.92). Among 3 eligible studies, no increased risk was observed for mortality (Peto OR 1.01; 95% CI 0.14-7.18) or infection (OR 0.58; 95% CI 0.27-1.27). The safety of lysine analogues in cancer patients has not been extensively studied. Based on the available literature, lysine analogue use has not been associated with increased risk of venous thromboembolism or other adverse events, while being effective in reducing blood loss and subsequent transfusion.

  17. A luminol chemiluminescence method for sensing histidine and lysine using enzyme reactions.

    PubMed

    Kugimiya, Akimitsu; Fukada, Rie; Funamoto, Daiki

    2013-12-01

    The analysis of free amino acids in urine and plasma is useful for estimating disease status in clinical diagnoses. Changes in the concentration of free amino acids in foods are also useful markers of freshness, nutrition, and taste. In this study, the specific interaction between aminoacyl-tRNA synthetase (aaRS) and its corresponding amino acid was used to measure amino acid concentrations. Pyrophosphate released by the amino acid-aaRS binding reaction was detected by luminol chemiluminescence; the method provided selective quantitation of 1.0-30 μM histidine and 1.0-60 μM lysine.

  18. Methionine one-electron oxidation: Coherent contributions from radiolysis, IRMPD spectroscopy, DFT calculations and electrochemistry

    NASA Astrophysics Data System (ADS)

    Scuderi, Debora; Bergès, Jacqueline; de Oliveira, Pedro; Houée-Levin, Chantal

    2016-11-01

    Methionine is an essential amino acid, unfortunately prone to oxidation. The mechanism of its oxidation by •OH radicals has been studied for more than 40 years and still remains misunderstood. We have reinvestigated the oxidation of this residue in model peptides, aiming at i) improving the identification of free radicals by the use of more modern quantum chemistry methods; ii) reinvestigating the one-electron reduction potentials as a function of the position in the sequence; iii) identifying the final compounds, which were still unknown; iv) reinvestigating the intramolecular electron transfer (IET) involving this residue.

  19. Transcriptional analysis of L-methionine catabolism in Brevibacterium linens ATCC9175.

    PubMed

    Cholet, Orianne; Hénaut, Alain; Bonnarme, Pascal

    2007-04-01

    The expression of genes possibly involved in L-methionine and lactate catabolic pathways were performed in Brevibacterium linens (ATCC9175) in the presence or absence of added L-methionine. The expression of 27 genes of 39 selected genes differed significantly in L-methionine-enriched cultures. The expression of the gene encoding L-methionine gamma-lyase (MGL) is high in L-methionine-enriched cultures and is accompanied by a dramatic increase in volatile sulfur compounds (VSC) biosynthesis. Several genes encoding alpha-ketoacid dehydrogenase and one gene encoding an acetolactate synthase were also up-regulated by L-methionine, and are probably involved in the catabolism of alpha-ketobutyrate, the primary degradation product of L-methionine to methanethiol. Gene expression profiles together with biochemical data were used to propose catabolic pathways for L-methionine in B. linens and their possible regulation by L-methionine.

  20. The impact of chronic imipramine treatment on amino acid concentrations in the hippocampus of mice.

    PubMed

    Nagasawa, Mao; Murakami, Tatsuro; Tomonaga, Shozo; Furuse, Mitsuhiro

    2012-09-01

    The relationship between antidepressants and monoamine concentrations in the brain has been well investigated, but few studies have investigated the relationship between antidepressants and amino acid concentrations in the brain. The purpose of the present study was therefore to investigate the effect of the chronic antidepressant imipramine on amino acid and monoamine concentrations in the mouse brain and plasma. Chronic imipramine treatment decreased the concentration of 5-hydroxyindoleaceticacid/5-hydroxytryptamine in the cerebral cortex and increased that of norepinephrine (NE) in the hippocampus. Since these changes were conspicuous effects of the antidepressant, we concluded that imipramine acts on the central nervous system. No change in amino acid concentrations in plasma was induced by chronic imipramine treatment, but several changes were confirmed in the cerebral cortex, the hypothalamus and the hippocampus. Chronic imipramine treatment caused increases in L-methionine, L-tyrosine, and L-lysine in the cerebral cortex, and an increase in L-aspartate in the hypothalamus. Contrary to this, the concentrations of L-aspartate, L-serine, L-asparagine, glycine, L-glutamine, gamma-aminobutyric acid, L-threonine, L-arginine, L-proline, L-valine, and L-methionine in the hippocampus were decreased by chronic imipramine treatment. The present results demonstrate that the metabolism of several amino acids in the brain, but not of those in plasma,